The transition zone protein Rpgrip1l regulates proteasomal activity at the primary cilium

Mutations in RPGRIP1L result in severe human diseases called ciliopathies. To unravel the molecular function of RPGRIP1L, we analyzed Rpgrip1l^−/− mouse embryos, which display a ciliopathy phenotype and die, at the latest, around birth. In these embryos, cilia-mediated signaling was severely disturbed. Defects in Shh signaling suggested that the Rpgrip1l deficiency causes an impairment of protein degradation and protein processing. Indeed, we detected a cilia-dependent decreased proteasomal activity in the absence of Rpgrip1l. We found different proteasomal components localized to cilia and identified Psmd2, a component of the regulatory proteasomal 19S subunit, as an interaction partner for Rpgrip1l. Quantifications of proteasomal substrates demonstrated that Rpgrip1l regulates proteasomal activity specifically at the basal body. Our study suggests that Rpgrip1l controls ciliary signaling by regulating the activity of the ciliary proteasome via Psmd2.     

Sensitivity Analysis of Environmental Process Modeling in a Life Cycle Context: A Case Study of Hemp Crop Production

The aim of this article is to develop a methodological approach allowing to assess the influence of parameters of one or more elementary processes in the foreground system, on the outcomes of a life cycle assessment (LCA) study. From this perspective, the method must be able to: (1) include foreground process modeling in order to avoid the assumption of proportionality between inventory data and reference flows; (2) quantify influences of foreground processes’ parameters (and, possibly, interactions between parameters); and (3) identify trends (either increasing or decreasing) for each parameter on each indicator in order to determine the most favorable direction for parametric variation. These objectives can be reached by combining foreground system modeling, a set of two different sensitivity analysis methods (each one providing different and complementary information), and LCA. The proposed method is applied to a case study of hemp‐based insulation materials for buildings. The present study will focus on the agricultural stage as a foreground system and as a first step encompassing the entire life cycle. A set of technological recommendations were identified for hemp farmers in order to reduce the crop's environmental impacts (from –11% to –89% according to the considered impact category). One of the main limitations of the approach is the need for a detailed model of the foreground process. Further, the method is, at present, rather time‐consuming. However, it offers long‐term advantages given that the higher level of model detail adds robustness to the LCA results.     

Fungal lysis by a soil bacterium fermenting cellulose

Recycling of plant biomass by a community of bacteria and fungi is fundamental to carbon flow in terrestrial ecosystems. Here we report how the plant fermenting, soil bacterium Clostridium phytofermentans enhances growth on cellulose by simultaneously lysing and consuming model fungi from soil. We investigate the mechanism of fungal lysis to show that among the dozens of different glycoside hydrolases C. phytofermentans secretes on cellulose, the most highly expressed enzymes degrade fungi rather than plant substrates. These enzymes, the GH18 Cphy1799 and Cphy1800, synergize to hydrolyse chitin, a main component of the fungal cell wall. Purified enzymes inhibit fungal growth and mutants lacking either GH18 grow normally on cellulose and other plant substrates, but have a reduced ability to hydrolyse chitinous substrates and fungal hyphae. Thus, C. phytofermentans boosts growth on cellulose by lysing fungi with its most highly expressed hydrolases, highlighting the importance of fungal interactions to the ecology of cellulolytic bacteria.     

Internalization of staphylococcal leukotoxins that bind and divert the C5a receptor is required for intracellular Ca2+ mobilization by human neutrophils

A growing number of receptors, often associated with the innate immune response, are being identified as targets for bacterial toxins of the beta‐stranded pore‐forming family. These findings raise the new question of whether the receptors are activated or merely used as docking points facilitating the formation of a pore. To elucidate whether the Staphylococcus aureus Panton‐Valentine leukocidin and the leukotoxin HlgC/HlgB act through the C5a receptor (C5aR) as agonists, antagonists or differ from the C5a complement‐derived peptide, their activity is explored on C5aR‐expressing cells. Both leukotoxins equally bound C5aR in neutrophils and in stable transfected U937 cells and initiated mobilization of intracellular Ca²⁺. HlgC/HlgB requires the presence of robust intracellular acidic Ca²⁺ stores in order to evoke a rise in free [Ca²⁺]_i, while the LukS‐PV/LukF‐PV directly altered reticular Ca²⁺ stores. Intracellular target specificity is conferred by the F‐subunit associated to the S‐subunit binding the receptor. Furthermore, internalization of the two leukotoxin components (S‐ and F‐subunits) associated to C5aR is required for the initiation of [Ca²⁺]_i mobilization. Electrophysiological recordings on living cells demonstrated that LukS‐PV/LukF‐PV does not alter the membrane resistance of C5aR‐expressing cells. The present observations suggest that part of the pore‐forming process occurs in distinct intracellular compartments rather than at the plasma membrane.     

The AvrE superfamily: ancestral type III effectors involved in suppression of pathogen‐associated molecular pattern‐triggered immunity

The AvrE superfamily of type III effectors (T3Es) is widespread among type III‐dependent phytobacteria and plays a crucial role during bacterial pathogenesis. Members of the AvrE superfamily are vertically inherited core effectors, indicating an ancestral acquisition of these effectors in bacterial plant pathogens. AvrE‐T3Es contribute significantly to virulence by suppressing pathogen‐associated molecular pattern (PAMP)‐triggered immunity. They inhibit salicylic acid‐mediated plant defences, interfere with vesicular trafficking and promote bacterial growth in planta. AvrE‐T3Es elicit cell death in both host and non‐host plants independent of any known plant resistance protein, suggesting an original interaction with the plant immune system. Recent studies in yeast have indicated that they activate protein phosphatase 2A and inhibit serine palmitoyl transferase, the first enzyme of the sphingolipid biosynthesis pathway. In this review, we describe the current picture that has emerged from studies of the different members of this fascinating large family.     

Coordination of cell proliferation and anterior-posterior axis establishment in the mouse embryo

During development, the growth of the embryo must be coupled to its patterning to ensure correct and timely morphogenesis. In the mouse embryo, migration of the anterior visceral endoderm (AVE) to the prospective anterior establishes the anterior-posterior (A-P) axis. By analysing the distribution of cells in S phase, M phase and G2 from the time just prior to the migration of the AVE until 18 hours after its movement, we show that there is no evidence for differential proliferation along the A-P axis of the mouse embryo. Rather, we have identified that as AVE movements are being initiated, the epiblast proliferates at a much higher rate than the visceral endoderm. We show that these high levels of proliferation in the epiblast are dependent on Nodal signalling and are required for A-P establishment, as blocking cell division in the epiblast inhibits AVE migration. Interestingly, inhibition of migration by blocking proliferation can be rescued by Dkk1. This suggests that the high levels of epiblast proliferation function to move the prospective AVE away from signals that are inhibitory to its migration. The finding that initiation of AVE movements requires a certain level of proliferation in the epiblast provides a mechanism whereby A-P axis development is coordinated with embryonic growth.     

Progressive Myoclonic Epilepsy-Associated Gene KCTD7 is a Regulator of Potassium Conductance in Neurons

The potassium channel tetramerization domain-containing protein 7 (KCTD7) was named after the structural homology of its predicted N-terminal broad complex, tramtrack and bric à brac/poxvirus and zinc finger domain with the T1 domain of the Kv potassium channel, but its expression profile and cellular function are still largely unknown. We have recently reported a homozygous nonsense mutation of KCTD7 in patients with a novel form of autosomal recessive progressive myoclonic epilepsy. Here, we show that KCTD7 expression hyperpolarizes the cell membrane and reduces the excitability of transfected neurons in patch clamp experiments. We found the expression of KCTD7 in the hippocampal and Purkinje cells of the murine brain, an expression profile consistent with our patients’ phenotype. The effect on the plasma membrane resting potential is possibly mediated by Cullin-3, as we demonstrated direct molecular interaction of KCTD7 with Cullin-3 in co-immunoprecipitation assays. Our data link progressive myoclonic epilepsy to an inherited defect of the neuron plasma membrane’s resting potential in the brain.