Person:
Meeske, Alexander Jacob

Profile Picture

Email Address

AA Acceptance Date

Birth Date

Research Projects

Organizational Units

Job Title

Last Name

Meeske

First Name

Alexander Jacob

Name

Meeske, Alexander Jacob
Author's Publications: search.filters.isAuthorOfPublication.ac651988-d410-4792-a0fa-e32bb94f458f

Search Results

Now showing 1 - 2 of 2
  • Thumbnail Image
    Publication
    High-Throughput Genetic Screens Identify a Large and Diverse Collection of New Sporulation Genes in Bacillus subtilis
    (Public Library of Science, 2016) Meeske, Alexander Jacob; Rodrigues, Christopher DA; Brady, Jacqueline; Lim, Hoong Chuin; Bernhardt, Thomas; Rudner, David
    The differentiation of the bacterium Bacillus subtilis into a dormant spore is among the most well-characterized developmental pathways in biology. Classical genetic screens performed over the past half century identified scores of factors involved in every step of this morphological process. More recently, transcriptional profiling uncovered additional sporulation-induced genes required for successful spore development. Here, we used transposon-sequencing (Tn-seq) to assess whether there were any sporulation genes left to be discovered. Our screen identified 133 out of the 148 genes with known sporulation defects. Surprisingly, we discovered 24 additional genes that had not been previously implicated in spore formation. To investigate their functions, we used fluorescence microscopy to survey early, middle, and late stages of differentiation of null mutants from the B. subtilis ordered knockout collection. This analysis identified mutants that are delayed in the initiation of sporulation, defective in membrane remodeling, and impaired in spore maturation. Several mutants had novel sporulation phenotypes. We performed in-depth characterization of two new factors that participate in cell–cell signaling pathways during sporulation. One (SpoIIT) functions in the activation of σE in the mother cell; the other (SpoIIIL) is required for σG activity in the forespore. Our analysis also revealed that as many as 36 sporulation-induced genes with no previously reported mutant phenotypes are required for timely spore maturation. Finally, we discovered a large set of transposon insertions that trigger premature initiation of sporulation. Our results highlight the power of Tn-seq for the discovery of new genes and novel pathways in sporulation and, combined with the recently completed null mutant collection, open the door for similar screens in other, less well-characterized processes.
  • No Thumbnail Available
    Publication
    Envelope biogenesis and spore formation in Bacillus subtilis
    (2016-05-02) Meeske, Alexander Jacob; Waldor, Matthew; Winston, Fred; Laub, Michael
    The bacterial cell wall exoskeleton (or peptidoglycan) protects cells from osmotic lysis, specifies their shape, and its biosynthetic pathway is a widely-exploited antibiotic target. My thesis research focused on defining how peptidoglycan synthesis is carried out in the model bacterium Bacillus subtilis. My first project explored the transport of cell wall precursors across the lipid bilayer. The precursor lipid II is a lipid-anchored molecule synthesized on the inner surface of the membrane. In E. coli, trans-bilayer movement of lipid II requires the essential flippase MurJ. I discovered that B. subtilis can survive in the absence of its 10 MurJ homologs through an alternate and novel lipid II flippase called Amj. I demonstrated that Amj is upregulated in response to envelope stress, suggesting that it serves as a defense mechanism against environmental stressors or antibiotics. My second project focused on the next step of the peptidoglycan biosynthesis pathway: the polymerization of lipid II into glycan strands. Polymerization is carried out by glycosyltransferases called class A penicillin-binding proteins (APBPs). In most bacteria, these enzymes are essential for viability, but they are dispensable in B. subtilis, implying the existence of an unidentified peptidoglycan synthase. I discovered that cells lacking these enzymes survive by upregulating a widely-conserved gene called rodA, whose homologs are essential for peptidoglycan assembly in virtually all bacteria. My biochemical analysis indicates that RodA has polymerase activity in vitro. Thus, B. subtilis, and likely most other bacteria, use two distinct mechanisms to synthesize their exoskeleton. RodA-like proteins represent appealing targets for broad-spectrum antibiotics. Inspired by high-throughput genetic screens being developed in the lab, I undertook a separate project focused on the developmental process of spore formation in B. subtilis. Genetic screens have identified factors with roles in every step of sporulation. Using a transposon sequencing approach, I identified 24 new sporulation genes, in addition to virtually all of the 148 previously characterized loci. Phenotypic characterization of these mutants uncovered factors involved in diverse aspects of sporulation, including a novel intercellular signaling molecule. My results highlight the power of transposon sequencing, and could inspire similar screens for other developmental processes.