| 1. |
- Panagiotou, Styliani, et al.
(författare)
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OSBP-mediated PI(4)P-cholesterol exchange at endoplasmic reticulum-secretory granule contact sites controls insulin secretion
- 2024
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Ingår i: Cell Reports. - : Cell Press. - 2211-1247. ; 43:4
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Tidskriftsartikel (refereegranskat)abstract
- Insulin is packaged into secretory granules that depart the Golgi and undergo a maturation process that involves changes in the protein and lipid composition of the granules. Here, we show that insulin secretory granules form physical contacts with the endoplasmic reticulum and that the lipid exchange protein oxysterol-binding protein (OSBP) is recruited to these sites in a Ca2+-dependent manner. OSBP binding to insulin granules is positively regulated by phosphatidylinositol-4 (PI4)-kinases and negatively regulated by the PI4 phosphate (PI(4)P) phosphatase Sac2. Loss of Sac2 results in excess accumulation of cholesterol on insulin granules that is normalized when OSBP expression is reduced, and both acute inhibition and small interfering RNA (siRNA)-mediated knockdown of OSBP suppress glucose-stimulated insulin secretion without affecting insulin production or intracellular Ca2+ signaling. In conclusion, we show that lipid exchange at endoplasmic reticulum (ER)-granule contact sites is involved in the exocytic process and propose that these contacts act as reaction centers with multimodal functions during insulin granule maturation.
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| 2. |
- Reimegård, Johan, et al.
(författare)
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Combined mRNA and protein single cell analysis in a dynamic cellular system
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Annan publikation (övrigt vetenskapligt/konstnärligt)abstract
- Combined measurements of mRNA and protein expression in single cells enables in-depth analysis of cellular states. We present single-cell protein and RNA co-profiling (SPARC), an approach to – for the first time – simultaneously measure global mRNA and large sets of intracellular protein in individual cells. Using SPARC, we show that mRNA expression fails to accurately reflect protein abundance at the time of measurement in human embryonic stem cells, although the direction of mRNA and protein expression changes is in agreement during cellular differentiation. Moreover, protein levels of transcription factors better predict their downstream effects than do their corresponding transcripts. We further show that changes of the balance between protein and mRNA expression levels can be applied to infer expression kinetic trajectories, predicting future states of individual cells. Finally, we highlight that protein expression variation is overall lower than mRNA variation, but relative variation of gene expression at the protein level does not reflect the mRNA level. Overall, our results demonstrate that mRNA and protein measurements in single cells provide different and complementary information regarding cell states. Accordingly, SPARC offers valuable insights into gene expression programs of single cells.
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| 3. |
- Reimegård, Johan, et al.
(författare)
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A combined approach for single-cell mRNA and intracellular protein expression analysis
- 2021
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Ingår i: Communications Biology. - : Springer Nature. - 2399-3642. ; 4:1
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Tidskriftsartikel (refereegranskat)abstract
- Combined measurements of mRNA and protein expression in single cells enable in-depth analysis of cellular states. We present SPARC, an approach that combines single-cell RNA-sequencing with proximity extension essays to simultaneously measure global mRNA and 89 intracellular proteins in individual cells. We show that mRNA expression fails to accurately reflect protein abundance at the time of measurement, although the direction of changes is in agreement during neuronal differentiation. Moreover, protein levels of transcription factors better predict their downstream effects than do their corresponding transcripts. Finally, we highlight that protein expression variation is overall lower than mRNA variation, but relative protein variation does not reflect the mRNA level. Our results demonstrate that mRNA and protein measurements in single cells provide different and complementary information regarding cell states. SPARC presents a state-of-the-art co-profiling method that overcomes current limitations in throughput and protein localization, including removing the need for cell fixation.
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| 4. |
- Xie, Beichen, et al.
(författare)
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The endoplasmic reticulum-plasma membrane tethering protein TMEM24 is a regulator of cellular Ca2+ homeostasis
- 2022
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Ingår i: Journal of Cell Science. - : The Company of Biologists. - 0021-9533 .- 1477-9137. ; 135:5
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Tidskriftsartikel (refereegranskat)abstract
- Endoplasmic reticulum (ER)-plasma membrane (PM) contacts are sites of lipid exchange and Ca2+ transport, and both lipid transport proteins and Ca2+ channels specifically accumulate at these locations. In pancreatic beta-cells, both lipid and Ca2+ signaling are essential for insulin secretion. The recently characterized lipid transfer protein TMEM24 (also known as C2CD2L) dynamically localizes to ER-PM contact sites and provides phosphatidylinositol, a precursor of phosphatidylinositol-4-phosphate [PI(4)P] and phosphatidylinositol 4,5-bisphosphate [PI(4,5)P-2], to the PM. beta-cells lacking TMEM24 exhibit markedly suppressed glucose-induced Ca2+ oscillations and insulin secretion, but the underlying mechanism is not known. We now show that TMEM24 onlyweakly interacts with the PM, and dissociates in response to both diacylglycerol and nanomolar elevations of cytosolic Ca2+. Loss of TMEM24 results in hyper-accumulation of Ca2+ in the ER and in excess Ca2+ entry into mitochondria, with resulting impairment in glucose-stimulated ATP production.
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