Host CDK-1 and formin mediate microvillar effacement induced by enterohemorrhagic Escherichia coli.

Enterohemorrhagic Escherichia coli (EHEC) induces changes to the intestinal cell cytoskeleton and formation of attaching and effacing lesions, characterized by the effacement of microvilli and then formation of actin pedestals to which the bacteria are tightly attached. Here, we use a Caenorhabditis elegans model of EHEC infection to show that microvillar effacement is mediated by a signalling pathway including mitotic cyclin-dependent kinase 1 (CDK1) and diaphanous-related formin 1 (CYK1). Similar observations are also made using EHEC-infected human intestinal cells in vitro. Our results support the use of C. elegans as a host model for studying attaching and effacing lesions in vivo, and reveal that the CDK1-formin signal axis is necessary for EHEC-induced microvillar effacement.

HLH-30/TFEB-mediated autophagy functions in a cell-autonomous manner for epithelium intrinsic cellular defense against bacterial pore-forming toxin in C. elegans.

Autophagy is an evolutionarily conserved intracellular system that maintains cellular homeostasis by degrading and recycling damaged cellular components. The transcription factor HLH-30/TFEB-mediated autophagy has been reported to regulate tolerance to bacterial infection, but less is known about the bona fide bacterial effector that activates HLH-30 and autophagy. Here, we reveal that bacterial membrane pore-forming toxin (PFT) induces autophagy in an HLH-30-dependent manner in Caenorhabditis elegans. Moreover, autophagy controls the susceptibility of animals to PFT toxicity through xenophagic degradation of PFT and repair of membrane-pore cell-autonomously in the PFT-targeted intestinal cells in C. elegans. These results demonstrate that autophagic pathways and autophagy are induced partly at the transcriptional level through HLH-30 activation and are required to protect metazoan upon PFT intoxication. Together, our data show a new and powerful connection between HLH-30-mediated autophagy and epithelium intrinsic cellular defense against the single most common mode of bacterial attack in vivo.

Biochemical characterization of the small ubiquitin-like modifiers of Chlamydomonas reinhardtii.

Dynamic modification of target proteins by small ubiquitin-like modifier (SUMO) is known to modulate many important cellular processes and is required for cell viability and development in all eukaryotes. However, understanding of SUMO systems in plants, especially in unicellular green algae, remains elusive. In this study, Chlamydomonas reinhardtii CrSUMO96, CrSUMO97 and CrSUMO148 were characterized. We show that the formation of polymeric CrSUMO96 and CrSUMO97 chains can be catalyzed either by the human SAE1/SAE2 and Ubc9 SUMOylation system in vitro or by an Escherichia coli chimeric SUMOylation system in vivo. An exposed C-terminal di-glycine motif of CrSUMO96 or CrSUMO97 is essential for functional SUMOylation. The human SUMO-specific protease, SENP1, demonstrates more processing activity for CrSUMO97 than for CrSUMO96. The CrSUMO148 precursor notably has four repeated di-glycine motifs at the C-terminus. This unique feature is not found in other known SUMO proteins. Interestingly, only 83-residual CrSUMO148(1-83) with the first di-glycine motif can form SAE1/SAE2-SUMO complex and further form polymeric chains with the help of Ubc9. More surprisingly, CrSUMO148 precursor is digested by SENP1, solely at the peptide bond after the first di-glycine motif although there are four theoretically identical processing sites in the primary sequence. This process directly generates 83-residual CrSUMO148(1-83) mature protein, which is exactly the form suitable for activation and conjugation. We also show that SENP1 displays similar isopeptidase activity in the deconjugation of polymeric CrSUMO96, CrSUMO97 or CrSUMO148 chains, revealing that the catalytic mechanisms of processing and deconjugation of CrSUMOs by SENP1 may differ.