Tailored approach to study Legionella infection using a lattice light sheet microscope (LLSM).

is a genus of ubiquitous environmental pathogens found in freshwater systems, moist soil, and composted materials. More than four decades of research has provided important insights into pathogenesis. Although standard commercial microscopes have led to significant advances in understanding pathogenesis, great potential exists in the deployment of more advanced imaging techniques to provide additional insights.

The UzcRS two-component system in Caulobacter crescentus integrates regulatory input from diverse auxiliary regulators.

The UzcRS two-component system in Caulobacter crescentus mediates widespread transcriptional activation in response to the metals U, Zn and Cu. Unexpectedly, a screen for mutations that affected the activity of the UzcR-regulated urcA promoter (P ) revealed four previously uncharacterized proteins whose inactivation led to metal-independent induction of P . Using molecular genetics and functional genomics, we find that these auxiliary regulators control P expression by modulating the activity of UzcRS through distinct mechanisms.

Identification of a U/Zn/Cu responsive global regulatory two-component system in Caulobacter crescentus.

Despite the well-known toxicity of uranium (U) to bacteria, little is known about how cells sense and respond to U. The recent finding of a U-specific stress response in Caulobacter crescentus has provided a foundation for studying the mechanisms of U- perception in bacteria. To gain insight into this process, we used a forward genetic screen to identify the regulatory components governing expression of the urcA promoter (P ) that is strongly induced by U.

Two Outer Membrane Proteins Contribute to Caulobacter crescentus Cellular Fitness by Preventing Intracellular S-Layer Protein Accumulation.

Surface layers, or S-layers, are two-dimensional protein arrays that form the outermost layer of many bacteria and archaea. They serve several functions, including physical protection of the cell from environmental threats. The high abundance of S-layer proteins necessitates a highly efficient export mechanism to transport the S-layer protein from the cytoplasm to the cell exterior.

Transposon Mutagenesis Paired with Deep Sequencing of Caulobacter crescentus under Uranium Stress Reveals Genes Essential for Detoxification and Stress Tolerance.

Unlabelled: The ubiquitous aquatic bacterium Caulobacter crescentus is highly resistant to uranium (U) and facilitates U biomineralization and thus holds promise as an agent of U bioremediation. To gain an understanding of how C. crescentus tolerates U, we employed transposon (Tn) mutagenesis paired with deep sequencing (Tn-seq) in a global screen for genomic elements required for U resistance.

Basal p21 controls population heterogeneity in cycling and quiescent cell cycle states.

Phenotypic heterogeneity within a population of genetically identical cells is emerging as a common theme in multiple biological systems, including human cell biology and cancer. Using live-cell imaging, flow cytometry, and kinetic modeling, we showed that two states--quiescence and cell cycling--can coexist within an isogenic population of human cells and resulted from low basal expression levels of p21, a Cyclin-dependent kinase (CDK) inhibitor (CKI). We attribute the p21-dependent heterogeneity in cell cycle activity to double-negative feedback regulation involving CDK2, p21, and E3 ubiquitin ligases.

The proliferation-quiescence decision is controlled by a bifurcation in CDK2 activity at mitotic exit.

Tissue homeostasis in metazoans is regulated by transitions of cells between quiescence and proliferation. The hallmark of proliferating populations is progression through the cell cycle, which is driven by cyclin-dependent kinase (CDK) activity. Here, we introduce a live-cell sensor for CDK2 activity and unexpectedly found that proliferating cells bifurcate into two populations as they exit mitosis.

Tuning gene expression with synthetic upstream open reading frames.

We engineered short ORFs and used them to control the expression level of recombinant proteins. These short ORFs, encoding a two-amino acid peptide, were placed upstream of an ORF encoding a protein of interest. Insertion of these upstream ORFs (uORFs) resulted in suppression of protein expression.

The Red clover necrotic mosaic virus capsid as a multifunctional cell targeting plant viral nanoparticle.

Multifunctional nanoparticles hold promise as the next generation of therapeutic delivery and imaging agents. Nanoparticles comprising many types of materials are being tested for this purpose, including plant viral capsids. It has been found that Red clover necrotic mosaic virus (RCNMV) can be loaded with significant amounts of therapeutic molecules with molecular weights of 600 or even greater.

Cellular uptake of gold nanoparticles passivated with BSA-SV40 large T antigen conjugates.

Internalization and subcellular localization in HeLa cells of gold nanoparticles modified with the SV40 large T antigen were quantified using inductively coupled plasma optical emission spectroscopy (ICP-OES). Internalization was monitored as a function of incubation time, temperature, nanoparticle diameter, and large T surface coverage. Increasing the amount of large T peptides per gold nanoparticle complex, by either increasing the coverage at constant nanoparticle diameter or by increasing the nanoparticle diameter at constant large T coverage, resulted in more cellular internalization.