Biology subjects

Hahn, J.

Publications and source records attributed to Hahn, J..

23 records · Page 2Linked to original sources

bioRxiv2021

Mechanisms of transforming DNA uptake to the periplasm of Bacillus subtilis

We demonstrate here that the acquisition of DNAase resistance by transforming DNA, often assumed to indicate transport to the cytoplasm, actually reflects uptake to the periplasm, requiring a re-evaluation of conclusions about the roles of several proteins in transformation. The new evidence suggests that the transformation pilus is needed for DNA binding to the cell surface near the cell poles and for the initiation of uptake. The cellular distribution of the membrane-anchored ComEA of B. subtilis does not noticeably change during DNA uptake as does the unanchored ComEA of Vibrio and Neisseria. Instead, our evidence suggests that ComEA stabilizes the attachment of transforming DNA at localized regions in the periplasm and then mediates uptake, probably by a Brownian ratchet mechanism. Following that, the DNA is transferred to periplasmic portions of the channel protein ComEC, which plays a previously unsuspected role in uptake to the periplasm. We show that the transformation endonuclease NucA also facilitates uptake to the periplasm and that the previously demonstrated role of ComFA in the acquisition of DNAase resistance actually derives from the instability of ComGA when ComFA is deleted. These results prompt a new understanding of the early stages of DNA uptake for transformation. IMPORTANCETransformation is a widely distributed mechanism of bacterial horizontal gene transfer that plays a role in the spread of antibiotic resistance and virulence genes and more generally in evolution. Although transformation was discovered nearly a century ago and most, if not all of the proteins required have been identified in several bacterial species, much remains poorly understood about the molecular mechanism of DNA uptake. This study uses epifluorescence microscopy to investigate the passage of labeled DNA into the compartment between the cell wall and the cell membrane of Bacillus subtilis, a necessary early step in transformation. The roles of individual proteins in this process are identified, and their modes of action are clarified.

microbiology↗

bioRxiv2020

ComEB protein is dispensable for transformation but must be translated for optimal synthesis of ComEC

We show that the ComEB protein is not required for transformation in Bacillus subtilis, despite its expression from within the comE operon under competence control. We show further that the synthesis of the putative channel protein ComEC is translationally coupled to the upstream comEB open reading frame, so that translation of comEB and a suboptimal ribosomal binding site embedded in its sequence are needed for proper comEC expression. Translational coupling appears to be a common mechanism in three major competence operons for the adjustment of protein amounts independent of transcriptional control, probably ensuring the correct stoichiometries for assembly of the transformation machinery. comEB and comFC respectively encode cytidine deaminase and a protein resembling type 1 phosphoribosyl transferases and we speculate that nucleotide scavenging proteins are produced under competence control for efficient reutilization of the products of degradation of the non-transforming strand during DNA uptake.

microbiology↗

bioRxiv2020

Molecular classification of zebrafish retinal ganglion cells links genes to cell types to behavior

Retinal ganglion cells (RGCs) form an array of feature detectors, which convey visual information to central brain regions. Characterizing RGC diversity is required to understand the logic of the underlying functional segregation. Using single-cell transcriptomics, we systematically classified RGCs in adult and larval zebrafish, thereby identifying marker genes for at least 33 stable and transient cell types. We used this dataset to engineer transgenic driver lines, enabling experimental access to specific RGC types. Strikingly, expression of one or few transcription factors often predicts dendrite morphologies and axonal projections to specific tectal layers and extratectal targets. In vivo calcium imaging revealed that molecularly defined RGCs exhibit highly specific functional tuning. Finally, chemogenetic ablation of eomesa+ RGCs, which comprise melanopsin-expressing types with projections to a small subset of central targets, selectively impaired phototaxis. Together, our study establishes a framework for systematically studying the functional architecture of the visual system.

neuroscience↗

bioRxiv2020

Sperm migration in the genital tract - in silico experiments identify key factors for reproductive success

Sperm migration in the female genital tract controls sperm selection and, therefore, reproductive success as male gametes are conditioned for fertilization while their number is dramatically reduced. Mechanisms underlying sperm migration are mostly unknown, since in vivo investigations are mostly unfeasible for ethical or practical reasons. By presenting a spatio-temporal model of the mammalian female genital tract combined with agent-based description of sperm motion and interaction as well as parameterizing it with bovine data, we offer an alternative possibility for studying sperm migration in silico. The model incorporates genital tract geometry as well as biophysical principles of sperm motion observed in vitro such as positive rheotaxis and thigmotaxis. This model for sperm migration from vagina to oviducts was successfully tested against in vivo data from literature. We found that physical sperm characteristics such as velocity and directional stability as well as sperm-fluid interactions and wall alignment are critical for success, i.e. sperms reaching the oviducts. Therefore, we propose that these identified sperm parameters should be considered in detail for conditioning sperm in artificial selection procedures since the natural processes are normally bypassed in reproductive in vitro technologies. The tremendous impact of mucus flow to support sperm accumulation in the oviduct highlights the importance of a species-specific optimum time window for artificial insemination regarding ovulation. Predictions from our extendable in silico experimental system will improve assisted reproduction in humans, endangered species, and livestock.

biophysics↗

bioRxiv2020

Genetic loci associated with prevalent and incident myocardial infarction and coronary heart disease in the Cohorts for Heart and Aging Research in Genomic Epidemiology (CHARGE) Consortium

BackgroundGenome-wide association studies have identified multiple genomic loci associated with coronary artery disease, but most are common variants in non-coding regions that provide limited information on causal genes and etiology of the disease. To better understand etiological pathways that might lead to discovery of new treatments or prevention strategies, we focused our investigation on low-frequency and rare sequence variations primarily residing in coding regions of the genome while also exploring associations with common variants. Methods and ResultsUsing samples of individuals of European ancestry from ten cohorts within the Cohorts for Heart and Aging Research in Genomic Epidemiology (CHARGE) consortium, both cross-sectional and prospective analyses were conducted to examine associations between genetic variants and myocardial infarction (MI), coronary heart disease (CHD), and all-cause mortality following these events. Single variant and gene-based analyses were performed separately in each cohort and then meta-analyzed for each outcome. A low-frequency intronic variant (rs988583) in PLCL1 was significantly associated with prevalent MI (OR=1.80, 95% confidence interval: 1.43, 2.27; P=7.12 x 10-7). Three common variants, rs9349379 in PHACTR1, and rs1333048 and rs4977574 in the 9p21 region, were significantly associated with prevalent CHD. Four common variants (rs4977574, rs10757278, rs1333049, and rs1333048) within the 9p21 locus were significantly associated with incident MI. We conducted gene-based burden tests for genes with a cumulative minor allele count (cMAC) [≥] 5 and variants with minor allele frequency (MAF) < 5%. TMPRSS5 and LDLRAD1 were significantly associated with prevalent MI and CHD, respectively, and RC3H2 and ANGPTL4 were significantly associated with incident MI and CHD, respectively. No loci were significantly associated with all-cause mortality following a MI or CHD event. ConclusionThis study confirmed previously reported loci influencing heart disease risk, and one single variant and three genes associated with MI and CHD were newly identified and warrant future investigation.

genetics↗