Currently: Consultant Chen, J. C., Hottes, A. K., McAdams, H. H., McGrath, P. T., Viollier, P. H., Shapiro, L. Two independent spiral structures control cell shape in Caulobacter. Biomedical Engineering, University of Toronto Quon, K. C., Yang, B., Domian, I. J., Shapiro, L., Marczynski, G. T. Transcriptional analysis of the Caulobacter 4.5 S RNA ffs gene and the physiological basis of an ffs mutant with a Ts phenotype, The CcrM DNA methyltransferase is widespread in the alpha subdivision of proteobacteria, and its essential functions are conserved in Rhizobium meliloti and Caulobacter crescentus, Identification of the fliI and fliJ components of the Caulobacter flagellar type III protein secretion system. centricity shift select prisma health; ontology and epistemology in nursing research; lamar county obituaries; nhs porter jobs glasgow; ottawa, ks police reports. Ramya Deshpande, SURF Scholar 2019-20 PhD at Harvard Analysis of mutant alleles of ctrA and point mutations in one of the CtrA binding sites in the origin demonstrate that CtrA represses replication in vivo. Homologs of CcrM are widespread in the alpha subdivision of the Proteobacteria, suggesting that methylation at GANTC sites may have important functions in other members of this diverse group as well. Dynamic protein localization is an integral component of the regulatory circuit that drives the Caulobacter cell cycle. RNA PRODUCT OF A REACTION CATALYZED BY A VIRAL RNA-DEPENDENT RNA POLYMERASE, Freeman Spogli Institute for International Studies, Institute for Computational and Mathematical Engineering (ICME), Institute for Human-Centered Artificial Intelligence (HAI), Institute for Stem Cell Biology and Regenerative Medicine, Stanford Institute for Economic Policy Research (SIEPR), Stanford Woods Institute for the Environment, Office of VP for University Human Resources, Office of Vice President for Business Affairs and Chief Financial Officer, Directed Reading in Developmental Biology, DOI 10.1146/annurev.genet.41.110306.130346, DOI 10.1146/annurev.biochem.72.121801.161824, DOI 10.1146/annurev.micro.56.012302.161103. A., Hillson, N. J., Shapiro, L. DipM links peptidoglycan remodelling to outer membrane organization in Caulobacter. Focused ultrasound excites cortical neurons by opening specific mechanosensitive ion channels, leading to gradual calcium accumulation, activation of calcium-gated channels, depolarization and spiking. An additional level of control was revealed when it was found that an interruption of DNA replication caused the inhibition of flaS transcription. These promoters, as well as those for several other genes encoding DNA replication proteins that are induced at the same time in the cell cycle, share two sequence motifs, suggesting that they represent a family whose transcription is coordinately regulated. Here we demonstrate live-cell 3D superresolution imaging of Crescentin-eYFP, a cytoskeletal fluorescent protein fusion, colocalized with the surface of the bacterium Caulobacter crescentus using a double-helix point spread function microscope. Thus, within the alpha subdivision, there is a conserved cell cycle dependence and regulatory mechanism controlling ccrM expression. As a result, we can selectively push them, trap them, pattern them and sort them selectively under acoustic remote control. These technologies take advantage of biomolecules with unusual physical properties allowing them to interact with sound waves and magnetic fields. The addition of rifampicin early after infection inhibited the production of phage, whereas phiCdl production was not inhibited by the addition of rifampicin at any time after infection of a rifampicin-resistant host. Because of the many parallels in the function of these biochemically based genetic circuits and electrical circuits, a hybrid modeling approach is proposed that integrates conventional biochemical kinetic modeling within the framework of a circuit simulation. Divergent region-specific regulation of neuronal excitability by muscarinic G. The cellular levels of CtrA and GcrA are temporally and spatially out-of-phase during the cell cycle, with CtrA repressing gcrA transcription and GcrA activating ctrA transcription. Strains with mutations in one of these genes, flaS, cannot transcribe flagellar structural genes and divide abnormally. Knowing a magnets past will allow scientists to customize particle beams more precisely in the future. View details for DOI 10.1146/annurev.genet.41.110306.130346, View details for Web of Science ID 000252359500018. Defects in the cheR gene resulted in a loss of the ability to methylate the methyl-accepting chemotaxis proteins. ErTadZ contains an atypical ATPase domain with a variant of a deviant Walker-A motif that retains ATP binding capacity while displaying only low intrinsic ATPase activity. We have carefully mapped out the multiple domains of KCNQ ion channels involved in interactions with phosphatidylinositol 4,5-bisphosphate (PIP2), a lipid signaling molecule of critical importance. Science Advances 9, eadd9186 (2023). Proteolytic control of Caulobacter cell cycle proteins is primarily executed by ClpXP, a dynamically localized protease implicated in turnover of several factors critical for faithful cell cycle progression. AN UNUSUAL PROMOTER CONTROLS CELL-CYCLE REGULATION AND DEPENDENCE ON DNA-REPLICATION OF THE CAULOBACTER-FLILM EARLY FLAGELLAR OPERON, PROTEIN LOCALIZATION AND ASYMMETRY IN THE BACTERIAL-CELL, FLOW-CYTOMETRY OF CAULOBACTER-CRESCENTUS - IDENTIFICATION AND CHARACTERIZATION OF A CELL-CYCLE MUTANT. 210-450-9060. Letts, V., Shaw, P., Shapiro, L., Henry, S. INVITRO TRANSCRIPTION OF THE EARLY REGION OF CAULOBACTER PHAGE PHI-CD1 DEOXYRIBONUCLEIC-ACID BY HOST RNA-POLYMERASE, 3-DIMENSIONAL RECONSTRUCTION OF THE FLAGELLAR HOOK FROM CAULOBACTER-CRESCENTUS. Spatial organization and dynamics of RNase E and ribosomes in Caulobacter crescentus. Ph.D. Student, Bioengineering (co-advised with Michael Elowitz) Learn more about the places where science happens at SLAC: our major facilities, institutes and centers. The biosynthesis of the single polar flagellum and the proteins that comprise the chemotaxis methylation machinery are both temporally and spacially regulated during the Caulobacter crescentus cell-division cycle. The research may lead to novel strategies for stimulating repair or regeneration of body tissues. The DnaA regulon includes genes encoding several replisome components, the GcrA global cell cycle regulator, the PodJ polar localization protein, the FtsZ cell division protein, and nucleotide biosynthesis enzymes. View details for DOI 10.1371/journal.pgen.1004463, View details for PubMedCentralID PMC4117421. In this study, we uncovered a mechanism by which daughter cell fate, which is specified by the DivJ-DivK-PleC system and effectively encoded in the phosphorylation state of the single-domain RR DivK, is communicated to the CckA-ChpT-CtrA signaling pathway that regulates more than 100 genes for polar differentiation, replication initiation and cell division. View details for Web of Science ID 000185536700040. PopZ interacts directly with the ParB protein bound to specific DNA sequences near the replication origin. Six distinct cellular characteristics, which are peculiar to these bacteria, have been defined and include (i) the synthesis of a polar organelle which may be membranous (21-23), (ii) a satellite DNA in the stalked cell (26), (iii) pili to which RNA bacteriophage specifically adsorb (16, 33), (iv) a single polar flagellum(17), (v) a lipopolysaccharide phage receptor site (27), and (vi) new cell wall material at the flagellated pole of the cell giving rise to a stalk (19, 20). During polar maturation, the PopZ-centromere tether is broken, and the PopZ zone at that pole then switches function to act as a recruitment factor for the ordered addition of multiple proteins that promote the transformation of the flagellated pole into a stalked pole. WebMikhail G. Shapiro, PhD Professor of Chemical Engineering & Medical Engineering Investigator, Howard Hughes Medical Institute Physics for Medicine Lab and Sorbonne Therefore, flagellar genes at or near the top of the hierarchy may be controlled, in part, by a unique transcription factor and may be responsive to the same DNA replication cues that mediate other cell cycle events, such as cell division. John Vaughen in Tom Clandinin lab successfully defended his thesis titled Sphingolipid Control of Neural Circuits by Glial Catabolism. Like Dam in the enterobacteria, CcrM plays a regulatory role in Caulobacter crescentus and Rhizobium meliloti. WebThe Leung Lab Research 1. Replisome assembly occurs at the chromosomal origin located at the stalked cell pole, coincident with the initiation of DNA replication. SURF Scholar 2022- View details for DOI 10.1073/pnas.2024705118, View details for Web of Science ID 000637394200069. In this review, we examine recently discovered control mechanisms that make use of dynamically localized protein complexes to orchestrate the Caulobacter crescentus cell cycle. x@caltech.edu, x=hsguo, Robert Hurt View details for Web of Science ID 000088048400024, View details for PubMedCentralID PMC16621. A peptide containing the C-terminal portion of the FtsA divisome protein is a substrate of both ClpXP and ClpAPin vitro but is primarily degraded by ClpAPin vivo. This control circuitry monitors the environment and the internal state of the cell, including the cell topology, as it orchestrates orderly activation of cell cycle subsystems and Caulobacter's asymmetric cell division. We discuss the genetic network and integrated three-dimensional sensor/response systems that regulate the cell cycle and asymmetric cell division in the bacterium Caulobacter crescentus. The chromosome is specifically and dynamically localized over the course of the cell cycle. View details for DOI 10.1016/j.molcel.2011.09.010, View details for Web of Science ID 000296212100011, View details for Web of Science ID 000299378306327. These discoveries have advanced our understanding of bacterial physiology and provided insight into the evolution of the eukaryotic cytoskeleton. Goley, E. D., Dye, N. A., Werner, J. N., Gitai, Z., Shapiro, L. CrfA, a small noncoding RNA regulator of adaptation to carbon starvation in Caulobacter crescentus. The effect of cyclic GMP derivatives was shown to be the repression of synthesis of specific structural proteins. Topologically-guided continuous protein crystallization controls bacterial surface layer self-assembly. Two phospho-signalling proteins, the transmembrane histidine kinase CckA and the cytoplasmic phosphotransferase ChpT, provide the only phosphate source for the cell fate-determining transcription factor CtrA9-18. Shapiro's laboratory question in developmental biology involves the mechanisms used to generate the three-dimensional organization of a cell from a one-dimensional genetic "New" poles generated at the cell division plane differ from old poles from the previous round of cell division. B.S. WebAs the first student, Soso declared open the lab at the 2016 Shriram new lab welcome party. View details for Web of Science ID 000083885400003. View details for DOI 10.1126/science.1095191, View details for Web of Science ID 000221383300040. The transcript synthesized in vitro was shown to be cleaved by C. crescentus RNase III and to release the transfer RNA genes from the downstream 16 S/23 S intergenic spacer region. M.Eng. Postdoctoral Scholar Previous studies have shown that flbT mutants overproduce flagellins and are unable to form chemotaxis swarm rings. Mutational analysis of FliI showed that two highly conserved amino acid residues in a bipartite ATP binding motif are necessary for flagellar assembly. View details for Web of Science ID 000294537800007, View details for PubMedCentralID PMC3202797. Two rings were in the hook-proximal upper set, and three rings (two narrow and one wide) were in the lower set. This type of gene overlap is also observed in bacterial genes involved in cell division. Future studies should integrate our knowledge of biochemical activities at Cori with our emerging understanding of cytological dynamics in C. crescentus and other bacteria. Ph.D. Engineering Science, University of Oxford x@caltech.edu, x=rohitk, Rohan Kolhe These Tn5 mutations had different effects on the methyl-accepting chemotaxis proteins (MCP), and the activities of methyltransferase and methylesterase. Quon, K. C., Marczynski, G. T., Shapiro, L. USE OF FLOW-CYTOMETRY TO IDENTIFY A CAULOBACTER 4.5 S RNA TEMPERATURE-SENSITIVE MUTANT DEFECTIVE IN THE CELL-CYCLE, A DEVELOPMENTALLY-REGULATED CHROMOSOMAL ORIGIN OF REPLICATION USES ESSENTIAL TRANSCRIPTION ELEMENTS. Postdoctoral Scholar, 2016-19 Structural maintenance of chromosomes proteins (SMCs) bind to DNA and function to ensure proper chromosome organization in both eukaryotes and bacteria. To study the relationship between phospholipid synthesis and organelle biogenesis in the dimorphic bacterium Caulobacter crescentus, auxotrophs have been isolated which require exogenous glycerol or glycerol 3-phosphate for growth when glucose is used as the carbon source. While not directly discernable, PopZ fills a region at the cell poles that is devoid of electron-dense ribosomes. The C. crescentus and Escherichia coli MCPs have highly conserved carboxy-terminal domains, and when an E. coli MCP is expressed in C. crescentus, it is targeted to the swarmer cell progeny. phiCbK DNA cosediments with Escherichia coli phage T2 DNA and has therefore been assigned an S(20,w) value of 63.5S. Many recent studies have revealed exquisite subcellular localization of proteins, DNA, and other molecules within bacterial cells, giving credence to the concept of prokaryotic anatomy. The analysis of small predivisional vesicles showed that the MCP content is higher in the flagellated vesicles, and analysis of large flagellated vesicles showed that the MCPs are positioned preferentially in the swarmer cell portion of the predivisional cell. Since many of these constructs are also suitable for use in other bacteria, this work provides a comprehensive collection of tools that will enrich many areas of microbiological research. (3,4) An additional global regulator, GcrA, has recently been discovered that both regulates and is regulated by CtrA. Therefore, we were able to recover a pH-conditional mutant in a cytoplasmic gene product. The hclA and the fatA genes mapped close together, possibly implying that comutation had occurred in AE6002. We also reexamined chromosome partitioning in a recombination-deficient strain of C. crescentus, and confirmed an earlier report that chromosomes partition to the progeny stalked and swarmer cells in a random manner that does not discriminate between old and new DNA strands. Caulobacter crescentus wild-type strain CB13 is unable to utilize galactose as the sole carbon source unless derivatives of cyclic AMP are present. Analysis of the requirements for CtrA polar localization and CtrA proteolysis revealed that both processes require a motif within amino acids 1-56 of the CtrA receiver domain, and neither process requires CtrA phosphorylation. View details for Web of Science ID A1997XV69900030, View details for PubMedCentralID PMC179479. The circuit diagram of the bacteriophage lambda lysislysogeny decision circuit represents connectivity in signal paths of the biochemical components. Phosphorylated CtrA (CtrA approximately P) bound to the CtrA recognition sequence in both promoters but had a 10- to 20-fold greater affinity for PfliQ. Cellular reproduction in all organisms requires temporal and spatial coordination of crucial events, notably DNA replication, chromosome segregation and cytokinesis. View details for DOI 10.1016/j.tcb.2007.03.005, View details for Web of Science ID 000246939100005. Mann, T. H., Seth Childers, W., Blair, J. University of Tehran In particular, little is known about the replication of multipartite genomes in bacteria. It is transcribed from three promoters; one is heat inducible, and the other two are induced at the transition from swarmer to stalked cell, coincident with the initiation of DNA replication. Shapiros lab has identified a pathway conserved from insects to humans, and between steroid hormones and peptide hormones in which mitogenic hormones, such as estrogen, and epidermal growth factor (EGF), elicit extremely rapid anticipatory activation of the stress sensor, the unfolded protein response (UPR). In bacteria, cell polarity has been observed by using both morphological and molecular markers; however, no general regulators of bacterial cell polarity have been identified. GapR interacts with adenine and thymine (AT)-rich chromosomal loci, associates with the promoter regions of cell cycle-regulated genes, and shares hundreds of recognition sites in common with known master regulators of cell cycle-dependent gene expression. x@caltech.edu, x=li.richard, Bill Ling Together, these results show that CcrM-catalyzed methylation adds another layer of control to the regulation of ctrA expression. Using purified protein in different oxidizing/reducing environments, we generate a calibration curve which can be used to analyze in situ measurements. Stanford AI Lab Papers and Talks at ICLR 2023. The Tn5 insertion mutant SC1130 had no cross-reacting MCP and had reduced levels of activity of the methyltransferase and methylesterase. This detailed beam information will help scientists perform their experiments more reliably a need that is becoming increasingly important as accelerator facilities operate at higher and higher energies and generate more complex beam profiles. SURF Scholar 2022 The Lon protease thus exhibits pleiotropic effects in C. crescentus growth and development. CtrA binds to and regulates the promoters of two genes critical to its temporally controlled proteolysis, divK and clpP, providing a transcriptional feedback loop for the control of cell cycle progression. The expression of the Caulobacter ccrM gene and the activity of its product, the M.Ccr II DNA methyltransferase, are limited to a discrete portion of the cell cycle (G. Zweiger, G. Marczynski, and L. Shapiro, J. Mol. The spatial distribution of the membrane chemoreceptors and associated cytoplasmic chemotaxis proteins in Escherichia coli were examined as a prototypic functional aggregate in bacterial cells. Genome analysis revealed that the C. crescentus genome encodes a significantly higher number of these signaling proteins (105) than any bacterial genome sequenced thus far. Removal of the membrane-spanning region of CckA results in loss of polar localization and cell death. The membrane-bound DivJ and PleC histidine kinases, which are asymmetrically localized at the opposite poles of the predivisional cell, control the temporal and spatial localization of DivK. The complete genome sequence of Caulobacter crescentus was determined to be 4,016,942 base pairs in a single circular chromosome encoding 3,767 genes. Eng. Lee, M., Schrader, J., Li, G., Weissman, J., McAdams, H., Shapiro, L., Moerner, W. E. DNA Segregation and Partitioning in Caulobacter Crescentus: Super-Resolving Protein Colocalization at the Cell Pole. The presence of SciP in the control network enhances the robustness of the cell cycle to varying growth rates. By analyzing mutations in the dnaX promoter, we identified a motif between the -10 and -35 regions that is required for proper timing of gene expression. We show that the S. meliloti CtrA belongs to the CtrA-like family of response regulators found in several alpha-proteobacteria. Several fla- mutants were also isolated by Tn5-VB32 mutagenesis and shown to confer kanamycin resistance. Here, we review the progress that has been made towards understanding the mechanisms by which bacterial cytoskeletal proteins influence cellular organization. Postdoctoral Scholar This project was supported in part by the DOEs Office of Science. An inducible promoter is a useful tool for the controlled expression of a given gene. Chemical Engineering, Cornell University Large structures, such as a flagellum, are anchored at the pole by means of the basal body that traverses the peptidoglycan wall. Bayas, C. A., Schrader, J. M., Lee, M. K., Shapiro, L., Moerner, W. E. CauloBrowser: A systems biology resource for Caulobacter crescentus. The chemoreceptor is localized at the flagellum-bearing pole of Caulobacter crescentus swarmer cells. Under such conditions the gene product itself, beta-galactosidase, is required to maintain intracellular pH, since such maintenance is clearly energy-dependent. Goley, E. D., Toro, E., McAdams, H. H., Shapiro, L. Superresolution Imaging in Live Caulobacter Crescentus Cells Using Photoswitchable Enhanced Yellow Fluorescent Protein, Super-resolution imaging in live Caulobacter crescentus cells using photoswitchable EYFP. Analysis of coliphage T7 in vitro transcripts showed that, like the E. coli enzyme, the C. crescentus RNA polymerase initiated transcription from the three major T7 early promoters and recognized the terminator at the end of the early region. The cell cycle-regulated methylation state of Caulobacter DNA mediates the temporal control of transcriptional activation of several key regulatory proteins. These new reporter genes provide much greater sensitivity, nonlinear ultrasound contrast, and ease-of-use for expression in a variety of cell types. Ph.D Biophysics (Imaging), University of Western Ontario Dynamic protease localization mediated by a phospho-signaling pathway is a novel mechanism to integrate spatial and temporal control of bacterial cell cycle progression. View details for DOI 10.1128/mBio.02238-16, View details for PubMedCentralID PMC5347347. Undergraduate Researcher 2022- Additional homologous sequences in phi X174 and a leader region of a ribosomal protein gene cluster were also detected. Such rotations did not lead to large-scale changes in gene expression, indicating that genome folding does not strongly affect gene regulation.
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