Biology subjects

Levin, P. A.

Publications and source records attributed to Levin, P. A..

5 recordsLinked to original sources

bioRxiv2018

Plasticity in Escherichia coli cell wall metabolism promotes fitness and mediates intrinsic antibiotic resistance across environmental conditions

Although the peptidoglycan cell wall is an essential structural and morphological feature of most bacterial cells, the extracytoplasmic enzymes involved in its synthesis are frequently dispensable under standard culture conditions. By modulating a single growth parameter--extracellular pH--we discovered a subset of these so-called \"redundant\" enzymes in Escherichia coli are required for maximal fitness across pH environments. Among these pH specialists are the class A penicillin binding proteins PBP1 a and PBP1 b; defects in these enzymes attenuate growth in alkaline and acidic conditions, respectively. Genetic, biochemical, and cytological studies demonstrate that synthase activity is required for cell wall integrity across a wide pH range, and differential activity across pH environments significantly alters intrinsic resistance to cell wall active antibiotics. Together, our findings reveal previously thought to be redundant enzymes are instead specialized for distinct environmental niches, thereby ensuring robust growth and cell wall integrity in a wide range of conditions.

microbiology↗

bioRxiv2018

Evolved sequence features within the intrinsically disordered tail influence FtsZ assembly and bacterial cell division

Intrinsically disordered regions (IDRs) challenge the well-established sequence-structure-function paradigm for describing protein function and evolution. Here, we direct a combination of biophysical and cellular studies to further our understanding of how the intrinsically disordered C-terminal tail of FtsZ contributes to cell division in rod-shaped bacteria. FtsZ is a modular protein that encompasses a conserved GTPase domain and a highly variable intrinsically disordered C-terminal tail (CTT). The CTT is essential for forming the cytokinetic Z-ring. Despite poor sequence conservation of the CTT, the patterning of oppositely charged residues, which refers to the extent of linear mixing / segregation of oppositely charged residues within CTT sequences is bounded within a narrow range. To assess the impact of evolutionary bounds on charge patterning within CTT sequences we performed experiments, aided by sequence design, to quantify the impact of changing the patterning of oppositely charged residues within the CTT on the functions of FtsZ from B. subtilis. Z-ring formation is robust if and only if the extent of linear mixing / segregation of oppositely charged residues within the CTT sequences is within evolutionarily observed bounds. Otherwise, aberrant, CTT-mediated, FtsZ assemblies impair Z-ring formation. The complexities of CTT sequences also have to be above a threshold value because FtsZ variants with low complexity CTTs are not tolerated in cells. Taken together, our results suggest that CTT sequences have evolved to be \"just right\" and that this is achieved through an optimal extent of charge patterning while maintaining the sequence complexity above a threshold value.

biophysics↗

bioRxiv2017

A nutrient-dependent division antagonist is regulated post-translationally by the Clp proteases in Bacillus subtilis

Changes in nutrient availability have dramatic and well-defined impacts on both transcription and translation in bacterial cells. At the same time, the role of post-translational control in adaptation to nutrient-poor environments is poorly understood. Here we report a role for the bacterial Clp proteases in degradation of the division inhibitor UgtP during growth in nutrient-poor medium. Under nutrient-rich conditions, interactions with its substrate UDP-glucose promote interactions between UgtP and the tubulin-like cell division protein FtsZ in Bacillus subtilis, inhibiting maturation of the cytokinetic ring and increasing cell size. In nutrient-poor medium, reductions in UDP-glucose availability favor UgtP oligomerization, sequestering it from FtsZ and allowing division to occur at a smaller cell mass. Intriguingly, in nutrient-poor conditions UgtP levels are reduced [~] 3-fold independent of UDP-glucose, suggesting an additional layer of regulation. B. subtilis cells cultured under different nutrient conditions indicate that UgtP accumulation is controlled through a nutrient-dependent post-translational mechanism dependent on the Clp proteases. Notably, all three B. subtilis Clp chaperones appeared able to target UgtP for degradation during growth in nutrient-poor conditions. Together these findings highlight conditional proteolysis as a mechanism for bacterial adaptation to a rapidly changing nutritional landscape.

microbiology↗

bioRxiv2016

Flux through lipid synthesis dictates bacterial cell size.

Nutrients--and by extension biosynthetic capacity--positively impact cell size in organisms throughout the tree of life. In bacteria, cell size is reduced three-fold in response to nutrient starvation or accumulation of the alarmone ppGpp, a global inhibitor of biosynthesis. However, whether biosynthetic capacity as a whole determines cell size or if particular anabolic pathways are more important than others remains an open question. Utilizing a top-down approach, here we identify flux through lipid synthesis as the primary biosynthetic determinant of Escherichia coli cell size. Altering flux through lipid synthesis recapitulated the impact of altering nutrients on cell size and morphology, while defects in other biosynthetic pathways either did not impact size or altered size in a lipid-dependent manner. Together our findings support a model in which lipid availability dictates cell envelope capacity and ppGpp functions as a linchpin linking surface area expansion with cytoplasmic volume to maintain cellular integrity.

microbiology↗

bioRxiv2016

High levels of antibiotic tolerance and persistence are induced by the commercial anti-microbial triclosan

The antimicrobial triclosan is used in a wide range of consumer products ranging from toothpaste, cleansers, socks, and baby toys. A bacteriostatic inhibitor of fatty acid synthesis, triclosan is extremely stable and accumulates in the environment. Approximately 75% of adults in the US have detectable levels of the compound in their urine, with a sizeable fraction of individuals (>10%) having urine concentrations equal to or greater than the minimal inhibitory concentration for Escherichia coli and methicillin-resistant Staphylococcus aureus (MRSA). Previous work has identified connections between defects in fatty acid synthesis and accumulation of the alarmone guanosine tetraphosphate (ppGpp), which has been repeatedly associated with antibiotic tolerance and persistence. Based on these data, we hypothesized that triclosan exposure may inadvertently drive bacteria into a state in which they are able to tolerate normally lethal concentrations of antibiotics. Here we report that clinically relevant concentrations of triclosan increased E. coli and MRSA tolerance to bactericidal antibiotics as much as 10,000 fold in vitro and reduced antibiotic efficacy up to 100-fold in a mouse urinary tract infection model. Genetic analysis indicated that triclosan-mediated antibiotic tolerance requires ppGpp synthesis, but is independent of growth. These data highlight an unexpected and certainly unintended consequence of adding high concentrations of antimicrobials in consumer products, supporting an urgent need to reevaluate the costs and benefits of the prophylactic use of triclosan and other bacteriostatic compounds.\n\nImportanceAdded as a prophylactic to a wide range of consumer products, the fatty acid synthesis inhibitor triclosan accumulates to high levels in humans and the environment. Based on links between defects in fatty acid synthesis and accumulation of the alarmone ppGpp, we hypothesized that triclosan would render cells tolerant to bactericidal compounds due to ppGpp-mediated inhibition of biosynthetic capacity. Our data indicate that clinically relevant concentrations of triclosan induces higher tolerance of E. coli and methicillin resistant S. aureus (MRSA) to a panel of bactericidal antibiotics up to 10,000-fold. In a urinary tract infection model, mice exposed to triclosan exhibited bacterial loads ~100-fold higher in the bladder than control animals following ciprofloxacin challenge. These findings highlight an unexpected consequence of antimicrobials in consumer products and support an urgent need to reevaluate the costs and benefits of the prophylactic use of triclosan and other bacteriostatic compounds.

microbiology↗