Biology subjects

Kroos, L.

Publications and source records attributed to Kroos, L..

3 recordsLinked to original sources

bioRxiv2023

Regulation of late-acting operons by three transcription factors and a CRISPR-Cas component during Myxococcus xanthus development

Upon starvation rod-shaped Myxococcus xanthus bacteria form mounds and then differentiate into round stress-resistant spores. Little is known about the regulation of late-acting operons important for spore formation. C-signaling has been proposed to activate FruA, which binds DNA cooperatively with MrpC to increase transcription of many genes. We report that this model can explain regulation of the fadIJ operon involved in spore metabolism, but not that of the spore coat biogenesis operons exoA-I, exoL-P, and nfsA-H. Rather, a mutation in fruA increased the transcript levels from these operons early in development, suggesting negative regulation by FruA initially, and a mutation in mrpC affected transcript levels from each operon differently. FruA bound to all four promoter regions in vitro, but strikingly each promoter region was unique in terms of whether or not MrpC and the DNA-binding domain of Nla6 bound, and in terms of cooperative binding. Furthermore, the DevI component of a CRISPR-Cas system is a negative regulator of all four operons, based on transcript measurements. Our results demonstrate complex regulation of sporulation genes by three transcription factors and a CRISPR-Cas component, which we propose thwarts viral intrusion while making spores suited to withstand starvation and environmental insults.

molecular biology↗

bioRxiv2021

Inhibitory Proteins Block Substrate Access to the Active Site of Bacillus subtilis Intramembrane Metalloprotease SpoIVFB

Intramembrane proteases function in numerous signaling pathways that impact health, but knowledge about regulation of intramembrane proteolysis is limited. We examined inhibition of intramembrane metalloprotease SpoIVFB by proteins BofA and SpoIVFA. We found that BofA residues in and near a predicted transmembrane segment are required for SpoIVFB inhibition, and cross-linking experiments indicated that this transmembrane segment occupies the SpoIVFB active site cleft. SpoIVFA is also required for SpoIVFB inhibition. The inhibitory proteins block access of the substrate N-terminal Proregion to the membrane-embedded SpoIVFB active site, based on additional cross-linking experiments; however, the inhibitory proteins did not prevent interaction between the substrate C-terminal region and the SpoIVFB soluble domain. A structural model was built of SpoIVFB in complex with BofA and parts of SpoIVFA and substrate, using partial homology and constraints from cross-linking and co-evolutionary analyses. The model predicts that conserved BofA residues interact to stabilize a transmembrane segment and a membrane-embedded C-terminal region. SpoIVFA is predicted to bridge the BofA C-terminal region and SpoIVFB, forming a membrane-embedded inhibition complex. Our results reveal a novel mechanism of intramembrane protease inhibition with clear implications for relief from inhibition in vivo and design of inhibitors as potential therapeutics.

biochemistry↗

bioRxiv2018

Systematic analysis of the Myxococcus xanthus developmental gene regulatory network supports posttranslational regulation of FruA by C-signaling

Upon starvation Myxococcus xanthus undergoes multicellular development. Rod-shaped cells move into mounds in which some cells differentiate into spores. Cells begin committing to sporulation at 24-30 h poststarvation, but the mechanisms governing commitment are unknown. FruA and MrpC are transcription factors that are necessary for commitment. They bind cooperatively to promoter regions and activate developmental gene transcription, including that of the dev operon. Leading up to and during the commitment period, dev mRNA increased in wild type, but not in a mutant defective in C-signaling, a short-range signaling interaction between cells that is also necessary for commitment. The C-signaling mutant exhibited [~]20-fold less dev mRNA than wild type at 30 h poststarvation, despite a similar level of MrpC and only twofold less FruA. Boosting the FruA level twofold in the C-signaling mutant had little effect on the dev mRNA level, and dev mRNA was not less stable in the C-signaling mutant. Neither did high cooperativity of MrpC and FruA binding upstream of the dev promoter explain the data. Rather, our systematic experimental and computational analyses support a model in which C-signaling activates FruA at least ninefold posttranslationally in order to commit a cell to spore formation.\n\nGraphical abstract\n\nO_FIG O_LINKSMALLFIG WIDTH=169 HEIGHT=200 SRC=\"FIGDIR/small/415331_ufig1.gif\" ALT=\"Figure 1\">\nView larger version (28K):\norg.highwire.dtl.DTLVardef@c16410org.highwire.dtl.DTLVardef@909252org.highwire.dtl.DTLVardef@17668a8org.highwire.dtl.DTLVardef@d126c3_HPS_FORMAT_FIGEXP M_FIG C_FIG Abbreviated summaryStarvation promotes MrpC accumulation, whereas nutrients favor proteolysis. MrpC activates transcription of fruA, but FruA protein appears to be activated by short-range C-signaling in a cycle leading to mound formation and lysis of some cells. Activated FruA* and MrpC are proposed to cooperatively stimulate transcription of the dev operon and genes that commit starving rod-shaped cells to form spores, while Dev proteins slow commitment, resulting in a spore-filled fruiting body surrounded by peripheral rods.

molecular biology↗