| 1. |
- Noborn, Fredrik, et al.
(författare)
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Role of neurexin heparan sulfate in the molecular assembly of synapses - Expanding the neurexin code?
- 2023
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Ingår i: Febs Journal. - : Wiley. - 1742-464X .- 1742-4658. ; 290:3, s. 252-265
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Tidskriftsartikel (refereegranskat)abstract
- Synapses are the minimal information processing units of the brain and come in many flavors across distinct circuits. The shape and properties of a synapse depend on its molecular organisation, which is thought to largely depend on interactions between cell adhesion molecules across the synaptic cleft. An established example is that of presynaptic neurexins and their interactions with structurally diverse postsynaptic ligands: the diversity of neurexin isoforms that arise from alternative promoters and alternative splicing specify synaptic properties by dictating ligand preference. The recent finding that a majority of neurexin isoforms exist as proteoglycans with a single heparan sulfate (HS) polysaccharide adds to this complexity. Sequence motifs within the HS polysaccharide may differ between neuronal cell types to contribute specificity to its interactions, thereby expanding the coding capacity of neurexin diversity. However, an expanding number of HS-binding proteins have been found capable to recruit neurexins via the HS chain, challenging the concept of a code provided by neurexin splice isoforms. Here we discuss the possible roles of the neurexin HS in light of what is known from other HS-protein interactions, and propose a model for how the neurexin HS polysaccharide may contribute to synaptic assembly. We also discuss how the neurexin HS may be regulated by co-secreted carbonic anhydrase-related and FAM19A proteins, and highlight some key issues that should be resolved to advance the field.
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| 2. |
- Uhlén, Mathias, et al.
(författare)
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A human protein atlas for normal and cancer tissues based on antibody proteomics
- 2005
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Ingår i: Molecular & Cellular Proteomics. - 1535-9476 .- 1535-9484. ; 4:12, s. 1920-1932
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Tidskriftsartikel (refereegranskat)abstract
- Antibody-based proteomics provides a powerful approach for the functional study of the human proteome involving the systematic generation of protein-specific affinity reagents. We used this strategy to construct a comprehensive, antibody-based protein atlas for expression and localization profiles in 48 normal human tissues and 20 different cancers. Here we report a new publicly available database containing, in the first version, similar to 400,000 high resolution images corresponding to more than 700 antibodies toward human proteins. Each image has been annotated by a certified pathologist to provide a knowledge base for functional studies and to allow queries about protein profiles in normal and disease tissues. Our results suggest it should be possible to extend this analysis to the majority of all human proteins thus providing a valuable tool for medical and biological research.
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| 3. |
- Ahmadian, Afshin, et al.
(författare)
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Single-nucleotide polymorphism analysis by pyrosequencing
- 2000
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Ingår i: Analytical Biochemistry. - : Elsevier BV. - 0003-2697 .- 1096-0309. ; 280:1, s. 103-110
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Tidskriftsartikel (refereegranskat)abstract
- There is a growing demand for high-throughput methods for analysis of single-nucleotide polymorphic (SNP) positions. Here, we have evaluated a novel sequencing approach, pyrosequencing, for such purposes. Pyrosequencing is a sequencing-by-synthesis method in which a cascade of enzymatic reactions yields detectable light, which is proportional to incorporated nucleotides. One feature of typing SNPs with pyrosequencing is that each allelic variant will give a unique sequence compared to the two other variants. These variants can easily be distinguished by a pattern recognition software. The software displays the allelic: alternatives and allows for direct comparison with the pyrosequencing raw data. For optimal determination of SNPs, various protocols of nucleotide dispensing order were investigated. Here, we demonstrate that typing of SNPs can efficiently be performed by pyrosequencing using an automated system for parallel analysis of 96 samples in approximately 5 min, suitable for large-scale screening and typing of SNPs.
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| 4. |
- Andersson, Anders, et al.
(författare)
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A transcriptional timetable of autumn senescence
- 2004
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Ingår i: Genome Biology. - : Springer Science and Business Media LLC. - 1465-6906 .- 1474-760X. ; 5:4, s. R24-
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Tidskriftsartikel (refereegranskat)abstract
- Background We have developed genomic tools to allow the genus Populus (aspens and cottonwoods) to be exploited as a full-featured model for investigating fundamental aspects of tree biology. We have undertaken large-scale expressed sequence tag (EST) sequencing programs and created Populus microarrays with significant gene coverage. One of the important aspects of plant biology that cannot be studied in annual plants is the gene activity involved in the induction of autumn leaf senescence. Results On the basis of 36,354 Populus ESTs, obtained from seven cDNA libraries, we have created a DNA microarray consisting of 13,490 clones, spotted in duplicate. Of these clones, 12,376 (92%) were confirmed by resequencing and all sequences were annotated and functionally classified. Here we have used the microarray to study transcript abundance in leaves of a free-growing aspen tree (Populus tremula) in northern Sweden during natural autumn senescence. Of the 13,490 spotted clones, 3,792 represented genes with significant expression in all leaf samples from the seven studied dates. Conclusions We observed a major shift in gene expression, coinciding with massive chlorophyll degradation, that reflected a shift from photosynthetic competence to energy generation by mitochondrial respiration, oxidation of fatty acids and nutrient mobilization. Autumn senescence had much in common with senescence in annual plants; for example many proteases were induced. We also found evidence for increased transcriptional activity before the appearance of visible signs of senescence, presumably preparing the leaf for degradation of its components.
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| 5. |
- Aspeborg, Henrik, et al.
(författare)
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Carbohydrate-active enzymes involved in the secondary cell wall biogenesis in hybrid aspen
- 2005
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Ingår i: Plant Physiology. - : Oxford University Press (OUP). - 0032-0889 .- 1532-2548. ; 137:3, s. 983-997
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Tidskriftsartikel (refereegranskat)abstract
- Wood formation is a fundamental biological process with significant economic interest. While lignin biosynthesis is currently relatively well understood, the pathways leading to the synthesis of the key structural carbohydrates in wood fibers remain obscure. We have used a functional genomics approach to identify enzymes involved in carbohydrate biosynthesis and remodeling during xylem development in the hybrid aspen Populus tremula x tremuloides. Microarrays containing cDNA clones from different tissue-specific libraries were hybridized with probes obtained from narrow tissue sections prepared by cryosectioning of the developing xylem. Bioinformatic analyses using the sensitive tools developed for carbohydrate-active enzymes allowed the identification of 25 xylem-specific glycosyltransferases belonging to the Carbohydrate-Active EnZYme families GT2, GT8, GT14, GT31, GT43, GT47, and GT61 and nine glycosidases (or transglycosidases) belonging to the Carbohydrate-Active EnZYme families GH9, GH10, GH16, GH17, GH19, GH28, GH35, and GH51. While no genes encoding either polysaccharide lyases or carbohydrate esterases were found among the secondary wall-specific genes, one putative O-acetyltransferase was identified. These wood-specific enzyme genes constitute a valuable resource for future development of engineered fibers with improved performance in different applications.
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| 6. |
- Bhalerao, Rupali, et al.
(författare)
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Gene expression in autumn leaves
- 2003
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Ingår i: Plant Physiology. - : Oxford University Press (OUP). - 0032-0889 .- 1532-2548. ; 131:2, s. 430-442
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Tidskriftsartikel (refereegranskat)abstract
- Two cDNA libraries were prepared, one from leaves of a field-grown aspen (Populus tremula) tree, harvested just before any visible sign of leaf senescence in the autumn, and one from young but fully expanded leaves of greenhouse-grown aspen (Populus tremula X tremuloides). Expressed sequence tags (ESTs; 5,128 and 4,841, respectively) were obtained from the two libraries. A semiautomatic method of annotation and functional classification of the ESTs, according to a modified Munich Institute of Protein Sequences classification scheme, was developed, utilizing information from three different databases. The patterns of gene expression in the two libraries were strikingly different. In the autumn leaf library, ESTs encoding metallothionein, early light-inducible proteins, and cysteine proteases were most abundant. Clones encoding other proteases and proteins involved in respiration and breakdown of lipids and pigments, as well as stress-related genes, were also well represented. We identified homologs to many known senescence-associated genes, as well as seven different genes encoding cysteine proteases, two encoding aspartic proteases, five encoding metallothioneins, and 35 additional genes that were up-regulated in autumn leaves. We also indirectly estimated the rate of plastid protein synthesis in the autumn leaves to be less that 10% of that in young leaves.
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| 7. |
- Djerbi, Soraya, et al.
(författare)
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The genome sequence of black cottonwood (Populus trichocarpa) reveals 18 conserved cellulose synthase (CesA) genes
- 2005
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Ingår i: Planta. - : Springer Science and Business Media LLC. - 0032-0935 .- 1432-2048. ; 221:5, s. 739-746
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Tidskriftsartikel (refereegranskat)abstract
- The genome sequence of Populus trichocarpa was screened for genes encoding cellulose synthases by using full-length cDNA sequences and ESTs previously identified in the tissue specific cDNA libraries of other poplars. The data obtained revealed 18 distinct CesA gene sequences in P. trichocarpa. The identified genes were grouped in seven gene pairs, one group of three sequences and one single gene. Evidence from gene expression studies of hybrid aspen suggests that both copies of at least one pair, CesA3-1 and CesA3-2, are actively transcribed. No sequences corresponding to the gene pair, CesA6-1 and CesA6-2, were found in Arabidopsis or hybrid aspen, while one homologous gene has been identified in the rice genome and an active transcript in Populus tremuloides. A phylogenetic analysis suggests that the CesA genes previously associated with secondary cell wall synthesis originate from a single ancestor gene and group in three distinct subgroups. The newly identified copies of CesA genes in P. trichocarpa give rise to a number of new questions concerning the mechanism of cellulose synthesis in trees.
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| 8. |
- Dujon, B, et al.
(författare)
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The nucleotide sequence of Saccharomyces cerevisiae chromosome XV
- 1997
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Ingår i: Nature. - : Springer Science and Business Media LLC. - 0028-0836 .- 1476-4687. ; 387:6632, s. 98-102
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Tidskriftsartikel (refereegranskat)abstract
- Chromosome XV was one of the last two chromosomes of Saccharomyces cerevisiae to be discovered(1). It is the third-largest yeast chromosome after chromosomes XII and IV, and is very similar in size to chromosome VII. It alone represents 9% of the yeast genome (8% if ribosomal DNA is included). When systematic sequencing of chromosome XV was started, 93 genes or markers were identified, and most of them were mapped(2). However, very little else was known about chromosome XV which, in contrast to shorter chromosomes, had not been the object of comprehensive genetic or molecular analysis. It was therefore decided to start sequencing chromosome XV only in the third phase of the European Yeast Genome Sequencing Programme, after experience was gained on chromosomes III, XI and II (refs 3-5). The sequence of chromosome XV has been determined from a set of partly overlapping cosmid clones derived from a unique yeast strain, and physically mapped at 3.3-kilobase resolution before sequencing. As well as numerous new open reading frames (ORFs) and genes encoding tRNA or small RNA molecules, the sequence of 1,091,283 base pairs confirms the high proportion of orphan genes and reveals a number of ancestral and successive duplications with other yeast chromosomes.
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| 9. |
- Finkelstein, D., et al.
(författare)
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Microarray data quality analysis : lessons from the AFGC project
- 2002
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Ingår i: Plant Molecular Biology. - 0167-4412 .- 1573-5028. ; 48:02-jan, s. 119-131
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Tidskriftsartikel (refereegranskat)abstract
- Genome-wide expression profiling with DNA microarrays has and will provide a great deal of data to the plant scientific community. However, reliability concerns have required the development data quality tests for common systematic biases. Fortunately, most large-scale systematic biases are detectable and some are correctable by normalization. Technical replication experiments and statistical surveys indicate that these biases vary widely in severity and appearance. As a result, no single normalization or correction method currently available is able to address all the issues. However, careful sequence selection, array design, experimental design and experimental annotation can substantially improve the quality and biological of microarray data. In this review, we discuss these issues with reference to examples from the Arabidopsis Functional Genomics Consortium (AFGC) microarray project.
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| 10. |
- Gray-Mitsumune, Madoka, et al.
(författare)
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Expansins abundant in secondary xylem belong to subgroup a of the alpha-expansin gene family (1 w )
- 2004
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Ingår i: Plant Physiology. - : Oxford University Press (OUP). - 0032-0889 .- 1532-2548. ; 135:3, s. 1552-1564
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Tidskriftsartikel (refereegranskat)abstract
- Differentiation of xylem cells in dicotyledonous plants involves expansion of the radial primary cell walls and intrusive tip growth of cambial derivative cells prior to the deposition of a thick secondary wall essential for xylem function. Expansins are cell wall-residing proteins that have an ability to plasticize the cellulose-hemicellulose network of primary walls. We found expansin activity in proteins extracted from the cambial region of mature stems in a model tree species hybrid aspen (Populus tremula X Populus tremuloides Michx). We identified three a-expansin genes (PttEXP1, PttEXP2, and PttEXP8) and one beta-expansin gene (PttEXPB1) in a cambial region expressed sequence tag library, among which PttEXP1 was most abundantly represented. Northern-blot analyses in aspen vegetative organs and tissues showed that PttEXP1 was specifically expressed in mature stems exhibiting secondary growth, where it was present in the cambium and in the radial expansion zone. By contrast, PttEXP2 was mostly expressed in developing leaves. In situ reverse transcription-PCR provided evidence for accumulation of mRNA of PttEXP1 along with ribosomal rRNA at the tips of intrusively growing xylem fibers, suggesting that PttEXP1 protein has a role in intrusive tip growth. An examination of tension wood and leaf cDNA libraries identified another expansin, PttEXP5, very similar to PttEXP1, as the major expansin in developing tension wood, while PttEXP3 was the major expansin expressed in developing leaves. Comparative analysis of expansins expressed in woody stems in aspen, Arabidopsis, and pine showed that the most abundantly expressed expansins share sequence similarities, belonging to the subfamily A of alpha-expansins and having two conserved motifs at the beginning and end of the mature protein, RIPVG and KNFRV, respectively. This conservation suggests that these genes may share a specialized, not yet identified function.
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| 11. |
- Jennions, Elizabeth, et al.
(författare)
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TANGO2 deficiency as a cause of neurodevelopmental delay with indirect effects on mitochondrial energy metabolism
- 2019
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Ingår i: Journal of Inherited Metabolic Disease. - : Wiley. - 0141-8955 .- 1573-2665. ; 42:5, s. 898-908
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Tidskriftsartikel (refereegranskat)abstract
- Exome sequencing has recently identified mutations in the gene TANGO2 (transport and Golgi organization 2) as a cause of developmental delay associated with recurrent crises involving rhabdomyolysis, cardiac arrhythmias, and metabolic derangements. The disease is not well understood, in part as the cellular function and subcellular localization of the TANGO2 protein remain unknown. Furthermore, the clinical syndrome with its heterogeneity of symptoms, signs, and laboratory findings is still being defined. Here, we describe 11 new cases of TANGO2-related disease, confirming and further expanding the previously described clinical phenotype. Patients were homozygous or compound heterozygous for previously described exonic deletions or new frameshift, splice site, and missense mutations. All patients showed developmental delay with ataxia, dysarthria, intellectual disability, or signs of spastic diplegia. Of importance, we identify two subjects (aged 12 and 17 years) who have never experienced any overt episode of the catabolism-induced metabolic crises typical for the disease. Mitochondrial complex II activity was mildly reduced in patients investigated in association with crises but normal in other patients. In one deceased patient, post-mortem autopsy revealed heterotopic neurons in the cerebral white matter, indicating a possible role for TANGO2 in neuronal migration. Furthermore, we have addressed the subcellular localization of several alternative isoforms of TANGO2, none of which were mitochondrial but instead appeared to have a primarily cytoplasmic localization. Previously described aberrations in Golgi morphology were not observed in cultured skin fibroblasts.
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| 12. |
- Johansson, H., et al.
(författare)
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Molecular cloning and characterization of a cDNA encoding poplar UDP-glucose dehydrogenase, a key gene of hemicellulose/pectin formation
- 2002
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Ingår i: Biochimica et Biophysica Acta, Gene Structure and Expression. - 0167-4781 .- 1879-2634. ; 1576:02-jan, s. 53-58
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Tidskriftsartikel (refereegranskat)abstract
- Plant UDP-glucose dehydrogenase (UGDH) is an important enzyme in the formation of hemicellulose and pectin, the components of newly formed cell walls. A cDNA clone (Ugdh) corresponding to UGDH was isolated from a cDNA library prepared from cambial zone of poplar (Populus tremula x tremuloides). Within the 1824-nucleotide (nt)-long clone, an open reading frame encoded a protein of 481 amino acids (aa), with a calculated molecular weight of 53.1 kDa. The derived aa sequence showed 90% and 63% identity with UGDHs from soybean and bovine liver, respectively, and had highly conserved aa motifs believed to be of importance for nt binding and catalytic efficiency. In poplar, the Ugdh corresponds to one or two genes, as found by genomic Southern analysis. The gene was expressed predominantly in differentiating xylem and young leaves, with little expression in the phloem zone of the stem. The expression pattern matched that of UGDH protein, as found by immunoblotting. In leaves, the Ugdh expression was upregulated by a short-term feeding with sucrose, sorbitol and polyethylene glycol, and this effect was to some extent mimicked by light exposure. The data suggest that Ugdh is regulated via an osmoticum-dependent pathway, possibly related to the availability of osmotically active carbohydrate precursors to UDPglucose, a substrate of UGDH.
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| 13. |
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| 14. |
- Karpinska, B., et al.
(författare)
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MYB transcription factors are differentially expressed and regulated during secondary vascular tissue development in hybrid aspen
- 2004
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Ingår i: Plant Molecular Biology. - : Springer Science and Business Media LLC. - 0167-4412 .- 1573-5028. ; 56:2, s. 255-270
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Tidskriftsartikel (refereegranskat)abstract
- More than 120,000 poplar ESTs have been sequenced from 20 different cDNA libraries by the Swedish Centre for Tree Functional Genomics. We screened this EST collection for MYB transcription factors involved in secondary vascular tissue formation, and genes assigned as PttMYB3Ra, PttMYB4a and PttMYB21a were selected for further characterisation. Three MYB genes showed different expression patterns in various organs, tissues and stem sub-sections representing different developmental stages of vascular tissue formation. Furthermore, the analysis showed that PttMYB21a expression was much higher in secondary cell wall formation zone of xylem and phloem fibers than in other developmental zones. Transgenic hybrid aspen plants, expressing the 3'-part of the PttMYB21a gene in antisense orientation were generated to assess the function of PttMYB21a gene in vascular tissue formation and lignification. All transgenic lines showed reduced growth and had fewer internodes compared to the wild-type. The analysis of selected lines showed that acid soluble lignin present in the bark was higher in transgenic lines as compared to wild-type plants. Moreover a higher transcript level of caffeoyl-CoA 3-O-methyltransferase [CCoAOMT];EC2.1.1.104) was found in the phloem of the transgenic plants, suggesting that PttMYB21a might function as a transcriptional repressor.
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| 15. |
- Kim, H., et al.
(författare)
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Calsyntenin-3 interacts with both ?- and ?-neurexins in the regulation of excitatory synaptic innervation in specific Schaffer collateral pathways
- 2020
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Ingår i: Journal of Biological Chemistry. - 0021-9258. ; 295:27, s. 9244-9262
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Tidskriftsartikel (refereegranskat)abstract
- Calsyntenin-3 (Clstn3) is a postsynaptic adhesion molecule that induces presynaptic differentiation via presynaptic neurexins (Nrxns), but whether Nrxns directly bind to Clstn3 has been a matter of debate. Here, using LC?MS/MS?based protein analysis, confocal microscopy, RNAscope assays, and electrophysiological recordings, we show that ?-Nrxns directly interact via their LNS domain with Clstn3 and Clstn3 cadherin domains. Expression of splice site 4 (SS4) insert?positive ?-Nrxn variants, but not insert?negative variants, reversed the impaired Clstn3 synaptogenic activity observed in Nrxn-deficient neurons. Consistently, Clstn3 selectively formed complexes with SS4?positive Nrxnsin vivo. Neuron-specific Clstn3 deletion caused significant reductions in number of excitatory synaptic inputs. Moreover, expression of Clstn3 cadherin domains in CA1 neurons of Clstn3 conditional knockout mice rescued structural deficits in excitatory synapses, especially within the stratum radiatum layer. Collectively, our results suggest that Clstn3 links to SS4?positive Nrxns to induce presynaptic differentiation and orchestrate excitatory synapse development in specific hippocampal neural circuits, including Schaffer collateral afferents.
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| 16. |
- Lindskog, Mats, 1974-
(författare)
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Computational analyses of biological sequences -applications to antibody-based proteomics and gene family characterization
- 2005
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Doktorsavhandling (övrigt vetenskapligt/konstnärligt)abstract
- Following the completion of the human genome sequence, post-genomic efforts have shifted the focus towards the analysis of the encoded proteome. Several different systematic proteomics approaches have emerged, for instance, antibody-based proteomics initiatives, where antibodies are used to functionally explore the human proteome. One such effort is HPR (the Swedish Human Proteome Resource), where affinity-purified polyclonal antibodies are generated and subsequently used for protein expression and localization studies in normal and diseased tissues. The antibodies are directed towards protein fragments, PrESTs (Protein Epitope Signature Tags), which are selected based on criteria favourable in subsequent laboratory procedures.This thesis describes the development of novel software (Bishop) to facilitate the selection of proper protein fragments, as well as ensuring a high-throughput processing of selected target proteins. The majority of proteins were successfully processed by this approach, however, the design strategy resulted in a number ofnfall-outs. These proteins comprised alternative splice variants, as well as proteins exhibiting high sequence similarities to other human proteins. Alternative strategies were developed for processing of these proteins. The strategy for handling of alternative splice variants included the development of additional software and was validated by comparing the immunohistochemical staining patterns obtained with antibodies generated towards the same target protein. Processing of high sequence similarity proteins was enabled by assembling human proteins into clusters according to their pairwise sequence identities. Each cluster was represented by a single PrEST located in the region of the highest sequence similarity among all cluster members, thereby representing the entire cluster. This strategy was validated by identification of all proteins within a cluster using antibodies directed to such cluster specific PrESTs using Western blot analysis. In addition, the PrEST design success rates for more than 4,000 genes were evaluated.Several genomes other than human have been finished, currently more than 300 genomes are fully sequenced. Following the release of the tree model organism black cottonwood (Populus trichocarpa), a bioinformatic analysis identified unknown cellulose synthases (CesAs), and revealed a total of 18 CesA family members. These genes are thought to have arisen from several rounds of genome duplication. This number is significantly higher than previous studies performed in other plant genomes, which comprise only ten CesA family members in those genomes. Moreover, identification of corresponding orthologous ESTs belonging to the closely related hybrid aspen (P. tremula x tremuloides) for two pairs of CesAs suggest that they are actively transcribed. This indicates that a number of paralogs have preserved their functionalities following extensive genome duplication events in the tree’s evolutionary history.
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| 17. |
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| 18. |
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| 19. |
- Lindskog, Mats, et al.
(författare)
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Selection of protein epitopes for antibody production
- 2005
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Ingår i: BioTechniques. - 0736-6205 .- 1940-9818. ; 38:5, s. 723-727
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Tidskriftsartikel (refereegranskat)abstract
- Protein functional analysis in the post-genomic era is a huge task that has to be approached by different methods in parallel. The use of protein-specific antibodies in conjunction with tissue microarrays has proven to be one important technology. In this study, we present a strategy for the optimized design of protein subfragments for subsequent antibody production. The fragments are selected based on a principle of lowest sequence similarity to other human proteins, optimally to generate antibodies with high selectivity. Furthermore, the fragments should have properties optimized for efficient protein production in Escherichia coli. The strategy has been implemented in Bishop, which is a Java-based software enabling the high-throughput production of protein fragments. Bishop allows for the avoidance of certain restriction enzyme sites, transmembrane regions, and signal peptides. A Basic Local Alignment Search Tool (BLAST) scanning procedure permits the selection of fragments of a selected size with a minimal sequence similarity to other proteins. The software and the strategy were evaluated on a human test data set and verified to fulfill the requested criteria.
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| 20. |
- Marco de La Cruz, Berta, 1990, et al.
(författare)
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Liprin-α proteins are master regulators of human presynapse assembly
- 2024
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Ingår i: NATURE NEUROSCIENCE. - 1097-6256 .- 1546-1726. ; 27, s. 629-642
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Tidskriftsartikel (refereegranskat)abstract
- The formation of mammalian synapses entails the precise alignment of presynaptic release sites with postsynaptic receptors but how nascent cell-cell contacts translate into assembly of presynaptic specializations remains unclear. Guided by pioneering work in invertebrates, we hypothesized that in mammalian synapses, liprin-alpha proteins directly link trans-synaptic initial contacts to downstream steps. Here we show that, in human neurons lacking all four liprin-alpha isoforms, nascent synaptic contacts are formed but recruitment of active zone components and accumulation of synaptic vesicles is blocked, resulting in 'empty' boutons and loss of synaptic transmission. Interactions with presynaptic cell adhesion molecules of either the LAR-RPTP family or neurexins via CASK are required to localize liprin-alpha to nascent synaptic sites. Liprin-alpha subsequently recruits presynaptic components via a direct interaction with ELKS proteins. Thus, assembly of human presynaptic terminals is governed by a hierarchical sequence of events in which the recruitment of liprin-alpha proteins by presynaptic cell adhesion molecules is a critical initial step. This paper identifies the evolutionarily conserved liprin-alpha protein family as key mediators of presynaptic assembly in human neurons. Their recruitment to sites formed by contacting neurons is the critical initial step that triggers presynaptic differentiation.
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| 21. |
- Montoliu-Gaya, Laia, et al.
(författare)
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CA10 regulates neurexin heparan sulfate addition via a direct binding in the secretory pathway
- 2021
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Ingår i: Embo Reports. - : EMBO. - 1469-221X .- 1469-3178. ; 22:4
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Tidskriftsartikel (refereegranskat)abstract
- Neurexins are presynaptic adhesion molecules that shape the molecular composition of synapses. Diversification of neurexins in numerous isoforms is believed to confer synapse-specific properties by engaging with distinct ligands. For example, a subset of neurexin molecules carry a heparan sulfate (HS) glycosaminoglycan that controls ligand binding, but how this post-translational modification is controlled is not known. Here, we observe that CA10, a ligand to neurexin in the secretory pathway, regulates neurexin-HS formation. CA10 is exclusively found on non-HS neurexin and CA10 expressed in neurons is sufficient to suppress HS addition and attenuate ligand binding and synapse formation induced by ligands known to recruit HS. This effect is mediated by a direct interaction in the secretory pathway that blocks the primary step of HS biosynthesis: xylosylation of the serine residue. NMR reveals that CA10 engages residues on either side of the serine that can be HS-modified, suggesting that CA10 sterically blocks xylosyltransferase access in Golgi. These results suggest a mechanism for the regulation of HS on neurexins and exemplify a new mechanism to regulate site-specific glycosylations.
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| 22. |
- Nguyen, N, et al.
(författare)
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Chromosomal sequencing using a PCR-based biotin-capture method allowed isolation of the complete gene for the outer membrane protein A of Klebsiella pneumoniae
- 1998
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Ingår i: Gene. - 0378-1119 .- 1879-0038. ; 210:1, s. 93-101
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Tidskriftsartikel (refereegranskat)abstract
- By employing a novel biotin-and PCR-assisted capture method, which allows determination of unknown sequences on chromosomal DNA. the gene for the outer membrane protein A (OmpA) of Klebsiella pneumoniae has been isolated and sequenced to completion. The method involves linear amplification of DNA from a biotinylated primer annealing to a region with known sequence. After capture of the amplified single-stranded DNA on to paramagnetic beads, unspecifically annealing primers, i.e. arbitrary primers, were used to generate fragments with only partly determined nt sequences. The homology of the sequenced gene to ompAs of related bacteria is discussed. The ompA gene was assembled for intracellular expression in Escherichia coli, and two different fusion proteins were produced and recovered with good yields. The importance of the novel chromosomal sequencing method for gene isolation in general and the potential use of the OmpA fusion proteins are discussed. (C) 1998 Elsevier Science B.V.
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| 23. |
- Oldfors Hedberg, Carola, 1969, et al.
(författare)
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Ribonuclease inhibitor 1 (RNH1) deficiency cause congenital cataracts and global developmental delay with infection-induced psychomotor regression and anemia
- 2023
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Ingår i: European Journal of Human Genetics. - : Springer Science and Business Media LLC. - 1018-4813 .- 1476-5438.
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Tidskriftsartikel (refereegranskat)abstract
- Ribonuclease inhibitor 1, also known as angiogenin inhibitor 1, encoded by RNH1, is a ubiquitously expressed leucine-rich repeat protein, which is highly conserved in mammalian species. Inactivation of rnh1 in mice causes an embryonically lethal anemia, but the exact biological function of RNH1 in humans remains unknown and no human genetic disease has so far been associated with RNH1. Here, we describe a family with two out of seven siblings affected by a disease characterized by congenital cataract, global developmental delay, myopathy and psychomotor deterioration, seizures and periodic anemia associated with upper respiratory tract infections. A homozygous splice-site variant (c.615-2A > C) in RNH1 segregated with the disease. Sequencing of RNA derived from patient fibroblasts and cDNA analysis of skeletal muscle mRNA showed aberrant splicing with skipping of exon 7. Western blot analysis revealed a total lack of the RNH1 protein. Functional analysis revealed that patient fibroblasts were more sensitive to RNase A exposure, and this phenotype was reversed by transduction with a lentivirus expressing RNH1 to complement patient cells. Our results demonstrate that loss-of-function of RNH1 in humans is associated with a multiorgan developmental disease with recessive inheritance. It may be speculated that the infection-induced deterioration resulted from an increased susceptibility toward extracellular RNases and/or other inflammatory responses normally kept in place by the RNase inhibitor RNH1.
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| 24. |
- Parra Bravo, Celeste, et al.
(författare)
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Human iPSC 4R tauopathy model uncovers modifiers of tau propagation.
- 2024
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Ingår i: Cell. - 1097-4172. ; 187:10
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Tidskriftsartikel (refereegranskat)abstract
- Tauopathies are age-associated neurodegenerative diseases whose mechanistic underpinnings remain elusive, partially due to a lack of appropriate human models. Here, we engineered human induced pluripotent stem cell (hiPSC)-derived neuronal lines to express 4R Tau and 4R Tau carrying the P301S MAPT mutation when differentiated into neurons. 4R-P301S neurons display progressive Tau inclusions upon seeding with Tau fibrils and recapitulate features of tauopathy phenotypes including shared transcriptomic signatures, autophagic body accumulation, and reduced neuronal activity. A CRISPRi screen of genes associated with Tau pathobiology identified over 500 genetic modifiers of seeding-induced Tau propagation, including retromer VPS29 and genes in the UFMylation cascade. In progressive supranuclear palsy (PSP) and Alzheimer's Disease (AD) brains, the UFMylation cascade is altered in neurofibrillary-tangle-bearing neurons. Inhibiting the UFMylation cascade invitro and invivo suppressed seeding-induced Tau propagation. This model provides a robust platform to identify novel therapeutic strategies for 4R tauopathy.
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| 25. |
- Sofou, Kalliopi, et al.
(författare)
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Bi-allelic VPS16 variants limit HOPS/CORVET levels and cause a mucopolysaccharidosis-like disease.
- 2021
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Ingår i: EMBO molecular medicine. - : EMBO. - 1757-4684 .- 1757-4676. ; 13
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Tidskriftsartikel (refereegranskat)abstract
- Lysosomal storage diseases, including mucopolysaccharidoses, result from genetic defects that impair lysosomal catabolism. Here, we describe two patients from two independent families presenting with progressive psychomotor regression, delayed myelination, brain atrophy, neutropenia, skeletal abnormalities, and mucopolysaccharidosis-like dysmorphic features. Both patients were homozygous for the same intronic variant in VPS16, a gene encoding a subunit of the HOPS and CORVET complexes. The variant impaired normal mRNA splicing and led to an ~85% reduction in VPS16 protein levels in patient-derived fibroblasts. Levels of other HOPS/CORVET subunits, including VPS33A, were similarly reduced, but restored upon re-expression of VPS16. Patient-derived fibroblasts showed defects in the uptake and endosomal trafficking of transferrin as well as accumulation of autophagosomes and lysosomal compartments. Re-expression of VPS16 rescued the cellular phenotypes. Zebrafish with disrupted vps16 expression showed impaired development, reduced myelination, and a similar accumulation of lysosomes and autophagosomes in the brain, particularly in glia cells. This disorder resembles previously reported patients with mutations in VPS33A, thus expanding the family of mucopolysaccharidosis-like diseases that result from mutations in HOPS/CORVET subunits.
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