Development of higher rates of nondiabetic glomerulosclerosis (GS) in African Americans has been attributed to two coding sequence variants (G1 and G2) in the APOL1 gene. To date, the cellular function and the role of APOL1 variants (Vs) in GS are still unknown. In this study, we examined the effects of overexpressing wild-type (G0) and kidney disease risk variants (G1 and G2) of APOL1 in human podocytes using a lentivirus expression system. Interestingly, G0 inflicted podocyte injury only at a higher concentration; however, G1 and G2 promoted moderate podocyte injury at lower and higher concentrations. APOL1Vs expressing podocytes displayed diffuse distribution of both Lucifer yellow dye and cathepsin L as manifestations of enhanced lysosomal membrane permeability (LMP). Chloroquine attenuated the APOL1Vs-induced increase in podocyte injury, consistent with targeting lysosomes. The chloride channel blocker DIDS prevented APOL1Vs- induced injury, indicating a role for chloride influx in osmotic swelling of lysosomes. Direct exposure of noninfected podocytes with conditioned media from G1- and G2-expressing podocytes also induced injury, suggesting a contributory role of the secreted component of G1 and G2 as well. Adverse host factors (AHFs) such as hydrogen peroxide, hypoxia, TNF-α, and puromycin aminonucleoside augmented APOL1- and APOL1Vs-induced podocyte injury, while the effect of human immunodeficiency virus (HIV) on podocyte injury was overwhelming under conditions of APOLVs expression. We conclude that G0 and G1 and G2 APOL1 variants have the potential to induce podocyte injury in a manner which is further augmented by AHFs, with HIV infection being especially prominent.

The contribution of African ancestry to the risk of focal segmental glomerulosclerosis and chronic kidney disease has been partially explained by the recently described chromosome 22q variants in the gene apolipoprotein L1 (APOL1). The APOL1 variants appear at a high allele frequency in populations of West African ancestry as a result of apparent adaptive selection of the heterozygous state. Heterozygosity protects from infection with Trypanosoma brucei rhodesiense. This review will describe the role of the approaches in population genetics for the description of APOL1-associated nephropathies and draw inferences as to the biologic mechanisms from genetic epidemiology findings to date. Modifier loci can influence APOL1 risk for the development of kidney disease. 'Second hits', both viral and non-viral, may explain the discrepancy between the remarkably high odds ratios and the low lifetime risks of kidney disease in two allele carriers of APOL1 risk variants. Therapeutic strategies for APOL1-associated nephropathies will require the prevention and treatment of these 'second hits' and the development of drugs to protect the APOL1 downstream renal injury pathways.

The 26S proteasome is a large cytoplasmic protease that degrades polyubiquitinated proteins to short peptides in a processive manner. The proteasome 19S regulatory subcomplex tethers the target protein via its polyubiquitin adduct and unfolds the target polypeptide, which is then threaded into the proteolytic sitecontaining 20S subcomplex. Hul5 is a 19S subcomplex-associated ubiquitin ligase that elongates ubiquitin chains on proteasome-bound substrates. We isolated hul5⌬ as a mutation with which fusions of an unstable cyclin to stable reporter proteins accumulate as partially processed products. These products appear transiently in the wild type but are strongly stabilized in 19S ATPase mutants and in the hul5⌬ mutant, supporting a role for the ATPase subunits in the unfolding of proteasome substrates before insertion into the catalytic cavity and suggesting a role for Hul5 in the processive degradation of proteins that are stalled on the proteasome.Selective degradation of cellular short-lived or damaged proteins occurs via the ubiquitin-proteasome system (UPS). Substrates of the UPS are covalently conjugated to the polypeptide ubiquitin via a cascade of enzymatic reactions mediated by a ubiquitin-activating enzyme, ubiquitin-conjugating enzymes, and ubiquitin ligases (16). The product of this reaction cascade is a chain of ubiquitin adducts, typically based on an isopeptide bond between the ε-amino group of a lysine residue of the substrate and the C-terminal carboxyl group of ubiquitin and with subsequent ubiquitin molecules similarly bound to an internal lysine of the preceding ubiquitin adduct. Polyubiquitinated substrates are subsequently recognized and degraded by the 26S proteasome, a large multicatalytic proteinase consisting of a catalytic barrel-shaped 20S subcomplex and two regulatory 19S subcomplexes at either end. The 20S subcomplex carries three distinct proteolytic activities-a chymotrypsin, trypsin, and caspase-like activity-carried by three different protein subunits (15,22,33,48). Access to the catalytic sites, which are located inside the lumen of the hollow 20S barrel, require passage through two narrow openings at either end of the barrel (12). This access is controlled by the 19S subcomplexes, the roles of which include binding polyubiquitinated proteins, unfolding the target polypeptide via a chaperone-like activity in order to allow it to be threaded into the 20S lumen, and hydrolyzing the polyubiquitin isopeptide bonds in order to recycle the ubiquitin (reviewed in references 6, 36, and 42). By analogy with eubacterial ATP-requiring proteases (29, 41) and archaebacterial proteasomes (44,50

Pho85 cyclins (Pcls), activators of the yeast cyclin-dependent kinase (CDK) Pho85, belong together with the p35 activator of mammalian CDK5 to a distinct structural cyclin class. Different Pcls target Pho85 to distinct substrates. Pcl5 targets Pho85 specifically to Gcn4, a yeast transcription factor involved in the response to amino acid starvation, eventually causing the degradation of Gcn4. Pcl5 is itself highly unstable, an instability that was postulated to be important for regulation of Gcn4 degradation. We used hybrids between different Pcls to circumscribe the substrate recognition function to the core cyclin box domain of Pcl5. Furthermore, the cyclin hybrids revealed that Pcl5 degradation is uniquely dependent on two distinct degradation signals: one N-terminal and one C-terminal to the cyclin box domain. Whereas the C-terminal degradation signal is independent of Pho85, the N-terminal degradation signal requires phosphorylation of a specific threonine residue by the Pho85 molecule bound to the cyclin. This latter mode of degradation depends on the SCF ubiquitin ligase. Degradation of Pcl5 after self-catalyzed phosphorylation ensures that activity of the Pho85/ Pcl5 complex is self-limiting in vivo. We demonstrate the importance of this mechanism for the regulation of Gcn4 degradation and for cell growth under conditions of amino acid starvation.Cyclin-dependent kinases (CDKs) are central cellular regulators that are involved in cell proliferation (46), as well as other processes, e.g., regulation of transcription via the phosphorylation of RNA polymerase II (7). CDKs are activated by binding of an ancillary subunit, the cyclin (32, 44). The cyclins also participate in targeting the kinase to specific substrates (28, 55, 65; reviewed in reference 43). Cyclins were so named because of their cyclic appearance and disappearance from the cells, in correlation with cell cycle progression (16). Their rapid and coordinated disappearance depends on the ubiquitin system, with the anaphase-promoting complex/cyclosome ubiquitin ligase being responsible for targeting mitotic cyclins, and SCF-type ubiquitin ligases often being involved in the degradation of G1 cyclins (68). However, not all cyclins are directly involved in cell cycle regulation, and not all cyclins are unstable. Common to all cyclins is that they possess a region of moderate sequence conservation, the cyclin box. The cyclin box was shown to assume a typical five-helix structure (8, 32), with helix 5 contacting the CDK near its catalytic site and helix 1 facing away from the CDK (32).The yeast CDK Pho85, a regulator of cell morphogenesis and of several metabolic pathways, is structurally and functionally related to the mammalian kinase CDK5 (29, 48). Of 10 Pho85 cyclin (Pcl) paralogs identified in yeast (40), many were associated with the phosphorylation of specific target proteins

Skeletal muscles respond to environmental and physiological changes by varying their size, fiber type, and metabolic properties. P38 mitogen-activated protein kinase (MAPK) is one of several signaling pathways that drive the metabolic adaptation of skeletal muscle to exercise. p38 MAPK also participates in the development of pathological traits resulting from excessive caloric intake and obesity that cause metabolic syndrome and type 2 diabetes (T2D). Whereas p38 MAPK increases insulin-independent glucose uptake and oxidative metabolism in muscles during exercise, it contrastingly mediates insulin resistance and glucose intolerance during metabolic syndrome development. This article provides an overview of the apparent contradicting roles of p38 MAPK in the adaptation of skeletal muscles to exercise and to pathological conditions leading to glucose intolerance and T2D. Here, we focus on the involvement of p38 MAPK in glucose metabolism of skeletal muscle, and discuss the possibility of targeting this pathway to prevent the development of T2D.

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Cardiac ATF3 has a protective role in dampening the HFD-induced cardiac remodeling processes. ATF3 exerts both local and systemic effects related to T2D-induced cardiomyopathy. This study provides a strong relationship between heart remodeling processes and blood glucose homeostasis.

The activating transcription factor 3 (ATF3) and the c-Jun dimerization protein 2 (JDP2) are members of the basic leucine zipper (bZIP) family of transcription factors. These proteins share a high degree of homology and both can activate or repress transcription. Deficiency of either one of them in the non-cancer host cells was shown to reduce metastases. As ATF3 and JDP2 compensate each other’s function, we studied the double deficiency of ATF3 and JDP2 in the stromal tumor microenvironment. Here, we show that mice with ATF3 and JDP2 double deficiency (designated thereafter dKO) developed larger tumors with high vascular perfusion and increased cell proliferation rate compared to wild type (WT) mice. We further identify that the underlying mechanism involves tumor associated fibroblasts which secrete high levels of stromal cell-derived factor 1 (SDF-1) in dKO fibroblasts. SDF-1 depletion in dKO fibroblasts dampened tumor growth and blood vessel perfusion. Furthermore, ATF3 and JDP2 were found to regulate SDF-1 transcription and secretion in fibroblasts, a phenomenon that is potentiated in the presence of cancer cells. Collectively, our results suggest that ATF3 and JDP2 regulate the expression of essential tumor promoting factors expressed by fibroblasts within the tumor microenvironment, and thus restrain tumor growth.