29 research outputs found
Peroxidasin-mediated crosslinking of collagen IV is independent of NADPH oxidases
Collagen IV is a major component of the basement membrane in epithelial tissues. The NC1 domains of collagen IV protomers are covalently linked together through sulfilimine bonds, the formation of which is catalyzed by peroxidasin. Although hydrogen peroxide is essential for this reaction, the exact source of the oxidant remains elusive. Members of the NOX/DUOX NADPH oxidase family are specifically devoted to the production of superoxide and hydrogen peroxide. Our aim in this study was to find out if NADPH oxidases contribute in vivo to the formation of collagen IV sulfilimine crosslinks. We used multiple genetically modified in vivo model systems to provide a detailed assessment of this question. Our data indicate that in various peroxidasin-expressing tissues sulfilimine crosslinks between the NC1 domains of collagen IV can be readily detected in the absence of functioning NADPH oxidases. We also analyzed how subatmospheric oxygen levels influence the collagen IV network in collagen-producing cultured cells with rapid matrix turnover. We showed that collagen IV crosslinks remain intact even under strongly hypoxic conditions. Our hypothesis is that during collagen IV network formation PXDN cooperates with a NOX/DUOX-independent H2O2 source that is functional also at very low ambient oxygen levels. © 201
Rab protein evolution and the history of the eukaryotic endomembrane system
Spectacular increases in the quantity of sequence data genome have facilitated major advances in eukaryotic comparative genomics. By exploiting homology with classical model organisms, this makes possible predictions of pathways and cellular functions currently impossible to address in intractable organisms. Echoing realization that core metabolic processes were established very early following evolution of life on earth, it is now emerging that many eukaryotic cellular features, including the endomembrane system, are ancient and organized around near-universal principles. Rab proteins are key mediators of vesicle transport and specificity, and via the presence of multiple paralogues, alterations in interaction specificity and modification of pathways, contribute greatly to the evolution of complexity of membrane transport. Understanding system-level contributions of Rab proteins to evolutionary history provides insight into the multiple processes sculpting cellular transport pathways and the exciting challenges that we face in delving further into the origins of membrane trafficking specificity
The leucine-rich repeat domain of human peroxidasin 1 promotes binding to laminin in basement membranes
Phospholipid Transfer Protein Sec14 Is Required for Trafficking from Endosomes and Regulates Distinct trans-Golgi Export Pathways*
A protein known to regulate both lipid metabolism and vesicular transport
is the phosphatidylcholine/phosphatidylinositol transfer protein Sec14 of
Saccharomyces cerevisiae. Sec14 is thought to globally affect
secretion from the trans-Golgi. The results from a synthetic genetic
array screen for genes whose inactivation impaired growth of cells with a
temperature-sensitive SEC14 allele implied Sec14 regulates transport
into and out of the Golgi. This prompted us to examine the role of Sec14 in
various vesicular transport pathways. We determined that Sec14 function was
required for the route followed by Bgl2, whereas trafficking of other secreted
proteins, including Hsp150, Cts1, Scw4, Scw10, Exg1, Cis3, and Ygp1, still
occurred, indicating Sec14 regulates specific trans-Golgi export
pathways. Upon diminution of Sec14 function, the v-SNARE Snc1 accumulated in
endosomes and the trans-Golgi. Its accumulation in endosomes is
consistent with Sec14 being required for transport from endosomes to the
trans-Golgi. Sec14 was also required for trafficking of Ste3 and the
lipophilic dye FM4-64 from the plasma membrane to the vacuole at the level of
the endosome. The combined genetic and cell biology data are consistent with
regulation of endosome trafficking being a major role for Sec14. We further
determined that lipid ligand occupancy differentially regulates Sec14
functions