| 1. |
- Behle, Anna, et al.
(författare)
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Manipulation of topoisomerase expression inhibits cell division but not growth and reveals a distinctive promoter structure in Synechocystis
- 2022
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Ingår i: Nucleic Acids Research. - : Oxford University Press (OUP). - 0305-1048 .- 1362-4962. ; 50:22, s. 12790-12808
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Tidskriftsartikel (refereegranskat)abstract
- In cyanobacteria DNA supercoiling varies over the diurnal cycle and is integrated with temporal programs of transcription and replication. We manipulated DNA supercoiling in Synechocystis sp. PCC 6803 by CRISPRi-based knockdown of gyrase subunits and overexpression of topoisomerase I (TopoI). Cell division was blocked but cell growth continued in all strains. The small endogenous plasmids were only transiently relaxed, then became strongly supercoiled in the TopoI overexpression strain. Transcript abundances showed a pronounced 5'/3' gradient along transcription units, incl. the rRNA genes, in the gyrase knockdown strains. These observations are consistent with the basic tenets of the homeostasis and twin-domain models of supercoiling in bacteria. TopoI induction initially led to downregulation of G+C-rich and upregulation of A+T-rich genes. The transcriptional response quickly bifurcated into six groups which overlap with diurnally co-expressed gene groups. Each group shows distinct deviations from a common core promoter structure, where helically phased A-tracts are in phase with the transcription start site. Together, our data show that major co-expression groups (regulons) in Synechocystis all respond differentially to DNA supercoiling, and suggest to re-evaluate the long-standing question of the role of A-tracts in bacterial promoters.
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| 3. |
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| 4. |
- Fedriani, Rubén, 1991, et al.
(författare)
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The SOFIA Massive (SOMA) Star Formation Survey. IV. Isolated Protostars
- 2023
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Ingår i: Astrophysical Journal. - : American Astronomical Society. - 1538-4357 .- 0004-637X. ; 942:1
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Tidskriftsartikel (refereegranskat)abstract
- We present similar to 10-40 mu m SOFIA-FORCAST images of 11 isolated protostars as part of the SOFIA Massive (SOMA) Star Formation Survey, with this morphological classification based on 37 mu m imaging. We develop an automated method to define source aperture size using the gradient of its background-subtracted enclosed flux and apply this to build spectral energy distributions (SEDs). We fit the SEDs with radiative transfer models, developed within the framework of turbulent core accretion (TCA) theory, to estimate key protostellar properties. Here, we release the sedcreator python package that carries out these methods. The SEDs are generally well fitted by the TCA models, from which we infer initial core masses M ( c ) ranging from 20-430 M (circle dot), clump mass surface densities sigma(cl) similar to 0.3-1.7 g cm(-2), and current protostellar masses m (*) similar to 3-50 M (circle dot). From a uniform analysis of the 40 sources in the full SOMA survey to date, we find that massive protostars form across a wide range of clump mass surface density environments, placing constraints on theories that predict a minimum threshold sigma(cl) for massive star formation. However, the upper end of the m (*)-sigma(cl) distribution follows trends predicted by models of internal protostellar feedback that find greater star formation efficiency in higher sigma(cl) conditions. We also investigate protostellar far-IR variability by comparison with IRAS data, finding no significant variation over an similar to 40 yr baseline.
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| 5. |
- Kaczmarzyk, Danuta, et al.
(författare)
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Diversion of the long-chain acyl-ACP pool in Synechocystis to fatty alcohols through CRISPRi repression of the essential phosphate acyltransferase PlsX
- 2018
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Ingår i: Metabolic engineering. - : Academic Press. - 1096-7176 .- 1096-7184. ; 45, s. 59-66
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Tidskriftsartikel (refereegranskat)abstract
- Fatty alcohol production in Synechocystis sp. PCC 6803 was achieved through heterologous expression of the fatty acyl-CoA/ACP reductase Maqu2220 from the bacteria Marinobacter aquaeolei VT8 and the fatty acyl-ACP reductase DPW from the rice Oryza sativa. These platform strains became models for testing multiplex CRISPR-interference (CRISPRi) metabolic engineering strategies to both improve fatty alcohol production and to study membrane homeostasis. CRISPRi allowed partial repression of up to six genes simultaneously, each encoding enzymes of acyl-ACP-consuming pathways. We identified the essential phosphate acyltransferase enzyme PlsX (slr1510) as a key node in C18 fatty acyl-ACP consumption, repression of slr1510 increased octadecanol productivity threefold over the base strain and gave the highest specific titers reported for this host, 10.3 mg g−1 DCW. PlsX catalyzes the first committed step of phosphatidic acid synthesis, and has not been characterized in Synechocystis previously. We found that accumulation of fatty alcohols impaired growth, altered the membrane composition, and caused a build-up of reactive oxygen species.
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| 6. |
- Law, Chi Yan, 1990, et al.
(författare)
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Isolated Massive Star Formation in G28.20-0.05
- 2022
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Ingår i: Astrophysical Journal. - : American Astronomical Society. - 1538-4357 .- 0004-637X. ; 939:2
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Tidskriftsartikel (refereegranskat)abstract
- We report high-resolution 1.3 mm continuum and molecular line observations of the massive protostar G28.20-0.05 with Atacama Large Millimeter/submillimeter Array. The continuum image reveals a ring-like structure with 2000 au radius, similar to morphology seen in archival 1.3 cm Very Large Array observations. Based on its spectral index and associated H30α emission, this structure mainly traces ionized gas. However, there is evidence for ∼30 M ⊙ of dusty gas near the main millimeter continuum peak on one side of the ring, as well as in adjacent regions within 3000 au. A virial analysis on scales of ∼2000 au from hot core line emission yields a dynamical mass of ∼80 M ⊙. A strong velocity gradient in the H30α emission is evidence for a rotating, ionized disk wind, which drives a larger-scale molecular outflow. An infrared spectral energy distribution (SED) analysis indicates a current protostellar mass of m * ∼ 40 M ⊙ forming from a core with initial mass M c ∼ 300 M ⊙ in a clump with mass surface density of Σcl ∼ 0.8 g cm−2. Thus the SED and other properties of the system can be understood in the context of core accretion models. A structure-finding analysis on the larger-scale continuum image indicates G28.20-0.05 is forming in a relatively isolated environment, with no other concentrated sources, i.e., protostellar cores, above ∼1 M ⊙ found from ∼0.1 to 0.4 pc around the source. This implies that a massive star can form in relative isolation, and the dearth of other protostellar companions within the ∼1 pc environs is a strong constraint on massive star formation theories that predict the presence of a surrounding protocluster.
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| 7. |
- Liu, Mengyao, et al.
(författare)
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The SOFIA Massive (SOMA) Star Formation Survey. III. From Intermediate- to High-mass Protostars
- 2020
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Ingår i: Astrophysical Journal. - : American Astronomical Society. - 1538-4357 .- 0004-637X. ; 904:1
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Tidskriftsartikel (refereegranskat)abstract
- We present similar to 10-40 mm SOFIA-FORCAST images of 14 intermediate-mass protostar candidates as part of the SOFIA Massive (SOMA) Star Formation Survey. We build spectral energy distributions, also using archival Spitzer, Herschel, and IRAS data. We then fit the spectral energy distributions with radiative transfer models of Zhang & Tan, based on turbulent core accretion theory, to estimate key protostellar properties. With the addition of these intermediate-mass sources, based on average properties derived from SED fitting, SOMA protostars span luminosities from similar to 10(2) to 10(6) L-circle dot, current protostellar masses from similar to 0.5 to 35 M-circle dot, and ambient clump mass surface densities, Scl, from 0.1 to g cm(-2). A wide range of evolutionary states of the individual protostars and of the protocluster environments is also probed. We have also considered about 50 protostars identified in infrared dark clouds that are expected to be at the earliest stages of their evolution. With this global sample, most of the evolutionary stages of high- and intermediate-mass protostars are probed. The best-fitting models show no evidence that a threshold value of the protocluster clump mass surface density is required to form protostars up to similar to 25 M.. However, to form more massive protostars, there is tentative evidence that Sigma(cl) needs to be greater than or similar to 1 g cm(-2). We discuss how this is consistent with expectations from core accretion models that include internal feedback from the forming massive star.
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| 8. |
- Santos, Marina, et al.
(författare)
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CRISPRi as a Tool to Repress Multiple Copies of Extracellular Polymeric Substances (EPS)-Related Genes in the Cyanobacterium Synechocystis sp. PCC 6803
- 2021
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Ingår i: Life. - : MDPI AG. - 2075-1729. ; 11:11
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Tidskriftsartikel (refereegranskat)abstract
- The use of the versatile cyanobacterial extracellular polymeric substances (EPS) for biotechnological/biomedical applications implies an extensive knowledge of their biosynthetic pathways to improve/control polymer production yields and characteristics. The multiple copies of EPS-related genes, scattered throughout cyanobacterial genomes, adds another layer of complexity, making these studies challenging and time-consuming. Usually, this issue would be tackled by generating deletion mutants, a process that in cyanobacteria is also hindered by the polyploidy. Thus, the use of the CRISPRi multiplex system constitutes an efficient approach to addressing this redundancy. Here, three putative Synechocystis sp. PCC 6803 kpsM homologues (slr0977, slr2107, and sll0574) were repressed using this methodology. The characterization of the 3-sgRNA mutant in terms of fitness/growth and total carbohydrates, released and capsular polysaccharides, and its comparison with previously generated single knockout mutants pointed towards Slr0977 being the key KpsM player in Synechocystis EPS production. This work validates CRISPRi as a powerful tool to unravel cyanobacterial complex EPS biosynthetic pathways expediting this type of studies.
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| 9. |
- Shabestary, Kiyan, et al.
(författare)
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Targeted Repression of Essential Genes To Arrest Growth and Increase Carbon Partitioning and Biofuel Titers in Cyanobacteria
- 2018
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Ingår i: ACS Synthetic Biology. - : American Chemical Society (ACS). - 2161-5063. ; 7:7
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Tidskriftsartikel (refereegranskat)abstract
- Photoautotrophic production of fuels and chemicals by cyanobacteria typically gives lower volumetric productivities and titers than heterotrophic production. Cyanobacteria cultures become light limited above an optimal cell density, so that this substrate is not supplied to all cells sufficiently. Here, we investigate genetic strategies for a two-phase cultivation, where biofuel-producing Synechocystis cultures are limited to an optimal cell density through inducible CRISPR interference (CRISPRi) repression of cell growth. Fixed CO2 is diverted to ethanol or n-butanol. Among the most successful strategies was partial repression of citrate synthase gltA. Strong repression (>90%) of gitA at low culture densities increased carbon partitioning to n-butanol 5-fold relative to a nonrepression strain, but sacrificed volumetric productivity due to severe growth restriction. CO2 fixation continued for at least 3 days after growth was arrested. By targeting sgRNAs to different regions of the gitA gene, we could modulate GItA expression and carbon partitioning between growth and product to increase both specific and volumetric productivity. These growth arrest strategies can be useful for improving performance of other photoautotrophic processes.
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| 10. |
- Yang, Yao-Lun, et al.
(författare)
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Constraining the Infalling Envelope Models of Embedded Protostars: BHR 71 and Its Hot Corino
- 2020
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Ingår i: Astrophysical Journal. - : American Astronomical Society. - 1538-4357 .- 0004-637X. ; 891:1
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Tidskriftsartikel (refereegranskat)abstract
- The collapse of a protostellar envelope results in the growth of a protostar and the development of a protoplanetary disk, playing a critical role during the early stages of star formation. Characterizing the gas infall in the envelope constrains the dynamical models of star formation. We present unambiguous signatures of infall, probed by optically thick molecular lines, toward an isolated embedded protostar, BHR 71 IRS1. The three-dimensional radiative transfer calculations indicate that a slowly rotating infalling envelope model following the "inside-out" collapse reproduces the observations of both lines, as well as the low-velocity emission of the HCN line. The envelope has a model-derived ag lines, where outflows or a Keplerian disk may contribute. The ALMA observations serendipitously discover the emission of complex organic molecules (COMs) concentrated within a radius of 100 au, indicating that BHR 71 IRS1 harbors a hot corino. Eight species of COMs are identified, including CH3OH and CH3OCHO, along with H2CS, SO2 and HCN v(2) = 1. The emission of methyl formate and C-13-methanol shows a clear velocity gradient within a radius of 50 au, hinting at an unresolved Keplerian rotating disk.
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