Search

Human C2orf69 is an evolutionarily conserved gene whose function is unknown. Here, we report eight unrelated families from which 20 children presented with a fatal syndrome consisting of severe autoinflammation and progredient leukoencephalopathy with recurrent seizures; 12 of these subjects, whose DNA was available, segregated homozygous loss-of-function C2orf69 variants. C2ORF69 bears homology to esterase enzymes, and orthologs can be found in most eukaryotic genomes, including that of unicellular phytoplankton. We found that endogenous C2ORF69 (1) is loosely bound to mitochondria, (2) affects mitochondrial membrane potential and oxidative respiration in cultured neurons, and (3) controls the levels of the glycogen branching enzyme 1 (GBE1) consistent with a glycogen-storage-associated mitochondriopathy. We show that CRISPR-Cas9-mediated inactivation of zebrafish C2orf69 results in lethality by 8 months of age due to spontaneous epileptic seizures, which is preceded by persistent brain inflammation. Collectively, our results delineate an autoinflammatory Mendelian disorder of C2orf69 deficiency that disrupts the development/homeostasis of the immune and central nervous systems.

The glycoproteome, the ensemble of glycans and their carrier proteins, plays major roles in multicellular life by regulating cell interactions with their environment. How information is encoded into the glycome, in other words how glycosylation is modulated in response to signals, remains largely unclear. Glycosylation enzymes operate predominantly in the endoplasmic reticulum (ER) and Golgi, a highly compartmentalized membrane-bound environment. Recent work indicates that this compartmentalization is plastic and tightly regulated. For instance, specific signals can induce the relocation of O-glycosylation enzymes, GALNTs, from the Golgi to the ER, resulting in significant upregulation of O-glycosylation initiation. We have named this re-compartmentation process the 'GALA pathway'. GALA illustrates how membrane trafficking in the secretory pathway can regulate protein glycosylation and thus encode information in the glycome.

The unfolded protein response (UPR) mediates the adaptation of the secretory pathway (SP) to fluctuations in cellular protein demand or to environmental variations. Recently, drug screenings have confirmed the therapeutic potential of targeting the UPR in cancer models. However, the UPR may not be the only druggable target of the SP. Moreover, recent studies have revealed other contributions of the SP to cancer development. This article does not intend to describe the well-established implication of UPR signaling pathways in cancer cell life and cell decision, but rather aims at defining the concept of 'tumor cell secretory addiction', from molecular, cellular, and therapeutic perspectives. Furthermore, the implication of UPR modulations in this context will be discussed.

SummaryWnts are transported to the cell surface by the integral membrane protein WLS (also known as Wntless, Evi, and GPR177). Previous studies of WLS trafficking have emphasized WLS movement from the Golgi to the plasma membrane (PM) and then back to the Golgi via retromer-mediated endocytic recycling. We find that endogenous WLS binds Wnts in the endoplasmic reticulum (ER), cycles to the PM, and then returns to the ER through the Golgi. We identify an ER-targeting sequence at the carboxyl terminus of native WLS that is critical for ER retrograde recycling and contributes to Wnt secretory function. Golgi-to-ER recycling of WLS requires the COPI regulator ARF as well as ERGIC2, an ER-Golgi intermediate compartment protein that is also required for the retrograde trafficking of the KDEL receptor and certain toxins. ERGIC2 is required for efficient Wnt secretion. ER retrieval is an integral part of the WLS transport cycle.

SummaryProtein toxins such as Ricin and Pseudomonas exotoxin (PE) pose major public health challenges. Both toxins depend on host cell machinery for internalization, retrograde trafficking from endosomes to the ER, and translocation to cytosol. Although both toxins follow a similar intracellular route, it is unknown how much they rely on the same genes. Here we conducted two genome-wide RNAi screens identifying genes required for intoxication and demonstrating that requirements are strikingly different between PE and Ricin, with only 13% overlap. Yet factors required by both toxins are present from the endosomes to the ER, and, at the morphological level, the toxins colocalize in multiple structures. Interestingly, Ricin, but not PE, depends on Golgi complex integrity and colocalizes significantly with a medial Golgi marker. Our data are consistent with two intertwined pathways converging and diverging at multiple points and reveal the complexity of retrograde membrane trafficking in mammalian cells.

O-GalNAc glycosylation of proteins confers essential structural, protective and signaling roles in eumetazoans. Addition of O-glycans onto proteins is an extremely complex process that regulates both sites of attachment and the types of oligosaccharides added. Twenty distinct polypeptide GalNAc-transferases (GalNAc-Ts) initiate O-glycosylation and fine-tuning their expression provides a mechanism for regulating this action. Recently, a new mode of regulation has emerged where activation of Src kinase selectively redistributes Golgi-localized GalNAc-Ts to the ER. This relocalization results in a strong increase in the density of O-glycan decoration. In this review, we discuss how different mechanisms can regulate the number and the types of O-glycans decorating proteins. In addition, we speculate how Src-dependent relocation of GalNAc-Ts could play an important role in cancerous cellular transformation.

Summary Cbl is an adaptor protein that is phosphorylated and recruited to several receptor and non-receptor tyrosine kinases upon their activation. After binding to the activated receptor, Cbl plays a key role as a kinase inhibitor and as an E3 ubiquitin ligase, thereby contributing to receptor down-regulation and internalization. In addition, Cbl translocates to intracellular vesicular compartments following receptor activation. We report here that Cbl also associates with Golgi membranes. Confocal immunofluorescence staining of Cbl in a variety of unstimulated cells, including CHO cells, revealed a prominent perinuclear colocalization of Cbl and a Golgi marker. Both the prominent Cbl staining and the Golgi marker were dispersed by brefeldin A. Subcellular fractionation of CHO cells demonstrated that about 10% of Cbl is stably associated with membranes, and that Golgi-enriched membrane fractions produced by isopycnic density centrifugation and free-flow electrophoresis are also enriched in Cbl, relative to other membrane fractions. The membrane-bound Cbl was hyperphosphorylated and it co-immunoprecipitated with endogenous Src. By immunofluorescence, some Src colocalized with Cbl and Golgi markers, and Src, like Cbl, was present in the Golgi-enriched fraction prepared by sequential density centrifugation and free-flow electrophoresis. Transfection of an activated form of Src, but not wild-type Src, increased the amount of Src that co-immunoprecipitated with Cbl, and increased the intensity of Cbl staining on the Golgi. This result, together with the increased tyrosine phosphorylation of the membrane-associated Cbl, suggests that Golgi-associated Cbl could be part of a molecular complex that contains activated Src. The localization and interaction of Src and Cbl at the Golgi and the regulation of the interaction of Cbl with Golgi membrane suggest that this complex may contribute to the regulation of Golgi function.

Summary Eur. J. Cell Biol. 81, 26–35 (2002). Upon request of the auhtors the name of Pierre Jurdic (Laboratoire de Biologie Moleculaire et Cellulaire, UMR 5665 CNRS/ENS, INRA, Ecole Normale Superieure de Lyon, Lyon/France) has to be added to the list of authors between J. B. Levy and W. C. Horne.