Engineering an Effective Human SNAP-23 Cleaving Botulinum Neurotoxin A Variant.

Botulinum neurotoxin (BoNT) serotype A inhibits neurotransmitter release by cleaving SNAP-25 and represents an established pharmaceutical for treating medical conditions caused by hyperactivity of cholinergic nerves. Oversecretion from non-neuronal cells is often also the cause of diseases. Notably, excessive release of inflammatory messengers is thought to contribute to diseases such as chronic obstructive pulmonary disease, asthma, diabetes etc.

Cut-and-Run: A Distinct Mechanism by which V(D)J Recombination Causes Genome Instability.

V(D)J recombination is essential to generate antigen receptor diversity but is also a potent cause of genome instability. Many chromosome alterations that result from aberrant V(D)J recombination involve breaks at single recombination signal sequences (RSSs). A long-standing question, however, is how such breaks occur.

ClpAP protease is a universal factor that activates the parDE toxin-antitoxin system from a broad host range RK2 plasmid.

The activity of type II toxin-antitoxin systems (TA), which are responsible for many important features of bacterial cells, is based on the differences between toxin and antitoxin stabilities. The antitoxin lability results from bacterial protease activity. Here, we investigated how particular Escherichia coli cytosolic proteases, namely, Lon, ClpAP, ClpXP, and ClpYQ, affect the stability of both the toxin and antitoxin components of the parDE system from the broad host range plasmid RK2.

Targeted genome regulation and modification using transcription activator-like effectors.

Transcription activator-like effectors (TALEs) are immensely powerful new tools for genome engineering that can be directed to bind to almost any DNA sequence of choice. They originate from the Xanthomonas species of plant pathogenic bacteria and, in nature, these proteins increase the virulence of Xanthomonas. However, in 2009, the DNA binding code of TALEs was deciphered and, subsequently, TALE proteins have been exploited for many diverse applications.

TALE proteins bind to both active and inactive chromatin.

TALE (transcription activator-like effector) proteins can be tailored to bind to any DNA sequence of choice and thus are of immense utility for genome editing and the specific delivery of transcription activators. However, to perform these functions, they need to occupy their sites in chromatin. In the present study, we have systematically assessed TALE binding to chromatin substrates and find that in vitro TALEs bind to their target site on nucleosomes at the more accessible entry/exit sites, but not at the nucleosome dyad.

Phosphorylation of the Smo tail is controlled by membrane localisation and is dispensable for clustering.

The Hedgehog (Hh) signalling cascade is highly conserved and involved in development and disease throughout evolution. Nevertheless, in comparison with other pathways, our mechanistic understanding of Hh signal transduction is remarkably incomplete. In the absence of ligand, the Hh receptor Patched (Ptc) represses the key signal transducer Smoothened (Smo) through an unknown mechanism.

Hh signalling is essential for somatic stem cell maintenance in the Drosophila testis niche.

In the Drosophila testis, germline stem cells (GSCs) and somatic cyst stem cells (CySCs) are arranged around a group of postmitotic somatic cells, termed the hub, which produce a variety of growth factors contributing to the niche microenvironment that regulates both stem cell pools. Here we show that CySC but not GSC maintenance requires Hedgehog (Hh) signalling in addition to Jak/Stat pathway activation. CySC clones unable to transduce the Hh signal are lost by differentiation, whereas pathway overactivation leads to an increase in proliferation.

Local BMP receptor activation at adherens junctions in the Drosophila germline stem cell niche.

According to the stem cell niche synapse hypothesis postulated for the mammalian haematopoietic system, spatial specificity of niche signals is maximized by subcellularly restricting signalling to cadherin-based adherens junctions between individual niche and stem cells. However, such a synapse has never been observed directly, in part, because tools to detect active growth factor receptors with subcellular resolution were not available. Here we describe a novel fluorescence-based reporter that directly visualizes bone morphogenetic protein (BMP) receptor activation and show that in the Drosophila testis a BMP niche signal is transmitted preferentially at adherens junctions between hub and germline stem cells, resembling the proposed synapse organization.

The Caenorhabditis elegans Ste20 kinase, GCK-3, is essential for postembryonic developmental timing and regulates meiotic chromosome segregation.

Ste20 kinases constitute a large family of serine/threonine kinases with a plethora of biological functions. Members of the GCK-VI subfamily have been identified as important regulators of osmohomeostasis across species functioning upstream of ion channels. Although the expression of the two highly similar mammalian GCK-VI kinases is eminent in a wide variety of tissues, which includes also the testis, their potential roles in development remain elusive.

GLS-1, a novel P granule component, modulates a network of conserved RNA regulators to influence germ cell fate decisions.

Post-transcriptional regulatory mechanisms are widely used to influence cell fate decisions in germ cells, early embryos, and neurons. Many conserved cytoplasmic RNA regulatory proteins associate with each other and assemble on target mRNAs, forming ribonucleoprotein (RNP) complexes, to control the mRNAs translational output. How these RNA regulatory networks are orchestrated during development to regulate cell fate decisions remains elusive.