Kathleen R Ryan

University of California, Berkeley | UCB · Department of Plant and Microbial Biology

Engineered living materials (ELMs) embed living cells in a biopolymer matrix to create materials with tailored functions. While bottom-up assembly of macroscopic ELMs with a de novo matrix would offer the greatest control over material properties, we lack the ability to genetically encode a protein matrix that leads to collective self-organization....

Lipid A, the membrane-anchored portion of lipopolysaccharide (LPS), is an essential component of the outer membrane (OM) of nearly all Gram-negative bacteria. Here we identify regulatory and structural factors that together render lipid A nonessential in Caulobacter crescentus. Mutations in the ferric uptake regulator fur allow Caulobacter to survi...

Caulobacter crescentus is an extremely informative model system for studies of the bacterial cell division cycle. Caulobacter resembles eukaryotes in that it performs chromosome replication once and only once per cell division. The ability to isolate G1-phase cells and examine them during synchronous passage through the division cycle has enabled s...

Lipid A, the membrane-anchored portion of lipopolysaccharide, is an essential component of the outer membrane (OM) of nearly all Gram-negative bacteria. Here, we identify regulatory and structural factors that together permit Caulobacter crescentus to eliminate lipid A from its OM. Mutations in the ferric uptake regulator fur allow Caulobacter to s...

Engineered living materials (ELMs) embed living cells in a biopolymer matrix to create novel materials with tailored functions. While bottom-up assembly of macroscopic ELMs with a de novo matrix would offer the greatest control over material properties, we lack the ability to genetically encode a protein matrix that leads to collective self-organiz...

The bacterial extracellular matrix forms autonomously, giving rise to complex material properties and multicellular behaviors. Synthetic matrix analogues can replicate these functions but require exogenously added material or have limited programmability. Here, we design a two-strain bacterial system that self-synthesizes and structures a synthetic...

The bacterial extracellular matrix forms autonomously, giving rise to complex material properties and multicellular behaviors. Synthetic matrix analogues can replicate these functions, but require exogenously added material or have limited programmability. Here we design a two-strain bacterial system that self-synthesizes and structures a synthetic...

Antibiotic persistence is a transient phenotypic state during which a bacterium can withstand otherwise lethal antibiotic exposure or environmental stresses. In Escherichia coli, persistence is promoted by the HipBA toxin-antitoxin system. The HipA toxin functions as a serine/threonine kinase that inhibits cell growth, while the HipB antitoxin neut...

Materials synthesized by organisms, such as bones and wood, combine the ability to self-repair with remarkable mechanical properties. This multifunctionality arises from the presence of living cells within the material and hierarchical assembly of different components across nanometer to micron scales. While creating engineered analogs of these nat...

Significance Controlled degradation of specific proteins is used by all organisms to change cell behavior in response to internal or external cues. Because ATP-dependent proteases, such as ClpXP, have a broad range of targets, accessory proteins called adaptors are often necessary for selective substrate proteolysis. Here we show that three protein...

How bacteria maintain their shape is not yet well understood. The bacterial cell wall is composed of peptidoglycan, which is a meshwork of glycan chains bonded covalently by short peptides. Since peptidoglycan is unique to bacteria, peptidoglycan synthesis proteins are well-known targets for antibiotic therapy against pathogenic bacteria. We want t...

The intricate cell division and developmental cycle of the α-Proteobacterium Caulobacter crescentus has been studied for four decades. During that time, elegant genetic screens and comprehensive post-genomic methods have uncovered a branched network of two-component signaling proteins that orchestrates Caulobacter cell cycle progression and morphol...

Although reversible phosphorylation on tyrosine residues regulates the activity of many eukaryotic proteins, there are few examples of this type of regulation in bacteria. We have identified the first essential tyrosine phosphatase homolog in a bacterium, Caulobacter crescentusCtpA. ctpAmutants with altered active-site residues are nonviable, and d...

The outer membrane of Gram-negative bacteria is an essential compartment containing a specific complement of lipids and proteins that constitute a protective, selective permeability barrier. Outer membrane beta-barrel proteins are assembled into the membrane by the essential hetero-oligomeric BAM complex, which contains the lipoprotein BamE. We hav...

Periodic activation and deactivation of the essential transcriptional regulator CtrA is necessary to drive cell cycle progression in Caulobacter crescentus. At the onset of DNA replication (the G1-S cell cycle transition), CtrA and the AAA+ protease ClpXP colocalize at one cell pole along with three accessory proteins, RcdA, CpdR, and PopA, and Ctr...

Recent studies have illuminated key features of the transcriptional and signal transduction networks governing the cell division cycle of Caulobacter crescentus. These mechanisms generate oscillations in the activity of CtrA, a key regulator of DNA replication and transcription of cell cycle regulated genes. ctrA transcription is itself regulated a...

The Caulobacter cell cycle is regulated by a network of two-component signal transduction proteins. Phosphorylation and stability of the master transcriptional regulator CtrA are controlled by the CckA-ChpT phosphorelay, and CckA activity is modulated by another response regulator, DivK. In a screen to identify suppressors of the cold-sensitive div...

How bacteria regulate cell cycle progression at a molecular level is a fundamental but poorly understood problem. In Caulobacter crescentus, two-component signal transduction proteins are crucial for cell cycle regulation, but the connectivity of regulators involved has remained elusive and key factors are unidentified. Here we identify ChpT, an es...

Regulated proteolysis is essential for cell cycle progression in both prokaryotes and eukaryotes. We show here that the ClpXP protease, responsible for the degradation of multiple bacterial proteins, is dynamically localized to specific cellular positions in Caulobacter where it degrades colocalized proteins. The CtrA cell cycle master regulator, t...

The first multicomponent phosphorelay, regulating stalk biogenesis, has been identified in Caulobacter crescentus using a bioinformatic screen, targeted disruptions of each histidine kinase and response regulator, and a new technique called phosphotransfer profiling, in which a purified histidine kinase or histidine phosphotransferase is simultaneo...

The response regulator CtrA, which silences the Caulobacter origin of replication and controls multiple cell cycle events, is specifically proteolyzed in cells preparing to initiate DNA replication. At the swarmer-to-stalked cell transition and in the stalked compartment of the predivisional cell, CtrA is localized to the cell pole just before its...

Asymmetric cell division in Caulobacter crescentus yields daughter cells that have different cell fates. Compartmentalization of the predivisional cell is a critical event in the establishment of the differential distribution of regulatory factors that specify cell fate. To determine when during the cell cycle the cytoplasm is compartmentalized so...

Bacteria exhibit a high degree of intracellular organization, both in the timing of essential processes and in the placement of the chromosome, the division site, and individual structural and regulatory proteins. We examine the temporal and spatial regulation of the Caulobacter cell cycle, bacterial chromosome segregation and cytokinesis, and Baci...

The two-component signaling protein CtrA activates or represses the expression of one-quarter of the cell-cycle-regulated genes in Caulobacter crescentus, integrating DNA replication, morphogenesis, and cell division. The activity of this essential protein is controlled by a positive transcriptional feedback loop, cell-cycle-regulated phosphorylati...

Clostridium perfringens perfringolysin O (PFO or theta-toxin) is a cytolytic toxin that binds to cholesterol-containing membranes and then self-associates to spontaneously form aqueous pores of varying size in the bilayer. In this study, a membrane-spanning domain has been identified in PFO by a combination of fluorescence spectroscopic methods usi...

The Tim23 protein is an essential inner membrane (IM) component of the yeast mitochondrial protein import pathway. Tim23p does not carry an amino-terminal presequence; therefore, the targeting information resides within the mature protein. Tim23p is anchored in the IM via four transmembrane segments and has two positively charged loops facing the m...

Tim23p is a mitochondrial inner membrane protein essential for the import of proteins from the cytosol. Tim23p contains an amino-terminal hydrophilic segment and a carboxyl-terminal hydrophobic domain (Tim23Cp). To study the functions and interactions of the two parts of Tim23p separately, we constructed tim23N , encoding only the hydrophilic regio...

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MAS6 encodes an essential inner membrane protein required for mitochondrial protein import in the yeast Saccharomyces cerevisiae (Emtage and Jensen, 1993). To identify new inner membrane import components, we isolated a high-copy suppressor (SMS1) of the mas6-1 mutant. SMS1 encodes a 16.5-kDa protein that contains several potential membrane-spannin...

Mas6p is an integral membrane protein of the yeast mitochondrial inner membrane, which is essential for mitochondrial protein import (1). To determine whether Mas6p is directly involved in recognizing precursors or translocating them across the inner membrane, we asked if Mas6p was in close proximity to precursor proteins being imported into mitoch...