Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:6.3.2.19 (ubiquitin-protein ligase)
 799 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

The epithelial Na+ channel (ENaC) was previously shown to be expressed in several Na(+)- and fluid-absorbing epithelia, particularly those of the kidney, colon, and lung. We have recently identified the ubiquitin-protein ligase Nedd4 as an interacting protein with ENaC and demonstrated that Nedd4 binds by its WW domains to the proline-rich PY motifs of ENaC. These PY motifs were recently shown to be deleted/mutated in patients afflicted with Liddle's syndrome, a hereditary form of systemic renal hypertension. Such mutations cause elevated channel activity by an increase in channel number/stability at the plasma membrane and by increased channel opening. We then proposed that Nedd4, by regulating channel stability/ degradation, may be a suppressor of ENaC. To test whether Nedd4 is localized to those tissues/regions that express ENaC, we performed immunocytochemical analysis of rat Nedd4 (rNedd4) distribution in rat kidney, colon, and lung tissues. Our results show that, in the kidney, rNedd4 is primarily localized to the cortical collecting tubules and outer and inner medullary collecting ducts. These tubular segments were previously shown to express ENaC. The epithelium lining medullary calyxes was also intensely stained, and microvillar borders of proximal convoluted tubules expressed variable amounts of rNedd4. In the lung, rNedd4 was mainly expressed in the epithelia lining the airways, in the submucosal glands and ducts, and in the distal respiratory epithelium. These sites resemble the pattern of ENaC expression. In contrast, in the distal colon, rNedd4 was strongly expressed in the epithelia lining the crypts but not in the ENaC-expressing surface epithelium. Low-salt diet (to elevate serum aldosterone levels) had no effect on rNedd4 distribution in the kidney or colon. Thus Nedd4 is coexpressed and likely colocalizes with ENaC in specific regions within the kidney and lung but not in the colon. We speculate this difference in colocalization may reflect differences in the regulation of channel stability in those tissues.
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PMID:Immunolocalization of the ubiquitin-protein ligase Nedd4 in tissues expressing the epithelial Na+ channel (ENaC). 922 16

The epithelial Na(+) channel (ENaC) regulates Na(+) absorption in epithelial tissues including the lung, colon and sweat gland, and in the distal nephrons of the kidney. When Na(+)-channel function is disrupted, salt and water homoeostasis is affected. The cytoplasmic regions of the Na(+)-channel subunits provide binding sites for other proteins to interact with and potentially regulate Na(+)-channel activity. Previously we showed that a proline-rich region of the alpha subunit of the Na(+) channel bound to a protein of 116 kDa from human lung cells. Here we report the identification of this protein as human Nedd4, a ubiquitin-protein ligase that binds to the Na(+)-channel subunits via its WW domains. Further, we show that WW domains 2, 3 and 4 of human Nedd4 bind to the alpha, beta and gamma Na(+)-channel subunits but not to a mutated beta subunit. In addition, when co-expressed in Xenopus oocytes, human Nedd4 down-regulates Na(+)-channel activity.
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PMID:Human Nedd4 interacts with the human epithelial Na+ channel: WW3 but not WW1 binds to Na+-channel subunits. 1064 8

Latent membrane protein 2A (LMP2A) is expressed in latent Epstein-Barr virus (EBV) infection. LMP2A functions to downregulate B-cell signal transduction and viral reactivation from latency in EBV-immortalized B cells in vitro, and acts to provide B cells with both a survival and developmental signal in vivo. Identification of proteins associated with LMP2A is important for elucidation of the mechanism that LMP2A employs to regulate B-cell signal transduction and EBV latency. LMP2A is constitutively tyrosine phosphorylated and is associated with protein tyrosine kinases such as Lyn and Syk when specific LMP2A tyrosines are phosphorylated. The amino-terminal domain of LMP2A includes multiple proline-rich regions, which may provide binding sites for proteins containing SH3 or WW domains. In this study, we demonstrate that four cellular proteins bind specifically to two PPPPY (PY) motifs present within the LMP2A amino-terminal domain. Protein microsequence analysis determined that three of these proteins were AIP4, WWP2/AIP2, and Nedd4. All of these proteins are members of the Nedd4-like ubiquitin-protein ligases family and have conserved domains including the C2, WW, and ubiquitin-protein ligase domain. The mutation of both PY motifs completely abolished binding activity of these proteins to LMP2A and the interaction of AIP4 and WWP2 with LMP2A was confirmed in cell lines expressing LMP2A, WWP2, and AIP4. Furthermore, a reduction in the level of Lyn and the rapid turnover of LMP2A and Lyn were observed in LMP2A-expressing cells. These findings suggest that LMP2A recruits Nedd4-like ubiquitin-protein ligases and B-cell signal transduction molecules, resulting in the degradation of LMP2A and Lyn by a ubiquitin-dependent mechanism. This provides a new means by which LMP2A may modulate B-cell signal transduction.
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PMID:The Epstein-Barr virus latent membrane protein 2A PY motif recruits WW domain-containing ubiquitin-protein ligases. 1068 40

RSP5 is an essential gene in Saccharomyces cerevisiae and was recently shown to form a physical and functional complex with RNA polymerase II (RNA pol II). The amino-terminal half of Rsp5 consists of four domains: a C2 domain, which binds membrane phospholipids; and three WW domains, which are protein interaction modules that bind proline-rich ligands. The carboxyl-terminal half of Rsp5 contains a HECT (homologous to E6-AP carboxyl terminus) domain that catalytically ligates ubiquitin to proteins and functionally classifies Rsp5 as an E3 ubiquitin-protein ligase. The C2 and WW domains are presumed to act as membrane localization and substrate recognition modules, respectively. We report that the second (and possibly third) Rsp5 WW domain mediates binding to the carboxyl-terminal domain (CTD) of the RNA pol II large subunit. The CTD comprises a heptamer (YSPTSPS) repeated 26 times and a PXY core that is critical for interaction with a specific group of WW domains. An analysis of synthetic peptides revealed a minimal CTD sequence that is sufficient to bind to the second Rsp5 WW domain (Rsp5 WW2) in vitro and in yeast two-hybrid assays. Furthermore, we found that specific "imperfect" CTD repeats can form a complex with Rsp5 WW2. In addition, we have shown that phosphorylation of this minimal CTD sequence on serine, threonine and tyrosine residues acts as a negative regulator of the Rsp5 WW2-CTD interaction. In view of the recent data pertaining to phosphorylation-driven interactions between the RNA pol II CTD and the WW domain of Ess1/Pin1, we suggest that CTD dephosphorylation may be a prerequisite for targeted RNA pol II degradation.
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PMID:Rsp5 WW domains interact directly with the carboxyl-terminal domain of RNA polymerase II. 1078 4

SLI-1, a Caenorhabditis elegans homologue of the proto-oncogene product c-Cbl, is a negative regulator of LET-23-mediated vulval differentiation. Lack of SLI-1 activity can compensate for decreased function of the LET-23 epidermal growth factor receptor, the SEM-5 adaptor, but not the LET-60 RAS, suggesting that SLI-1 acts before RAS activation. SLI-1 and c-Cbl comprise an N-terminal region (termed SLI-1:N/Cbl-N, containing a four-helix bundle, an EF hand calcium-binding domain, and a divergent SH2 domain) followed by a RING finger domain and a proline-rich C-terminus. In a transgenic functional assay, the proline-rich C-terminal domain is not essential for sli-1(+) function. A protein lacking the SH2 and RING finger domains has no activity, but a chimeric protein with the SH2 and RING finger domains of SLI-1 replaced by the equivalent domains of c-Cbl has activity. The RING finger domain of c-Cbl has been shown recently to enhance ubiquitination of active RTKs by acting as an E3 ubiquitin-protein ligase. We find that the RING finger domain of SLI-1 is partially dispensable. Further, we identify an inhibitory tyrosine of LET-23 requiring sli-1(+) for its effects: removal of this tyrosine closely mimics the loss of sli-1 but not of another negative regulator, ark-1. Thus, we suggest that this inhibitory tyrosine mediates its effects through SLI-1, which in turn inhibits signaling upstream of LET-60 RAS in a manner not wholly dependent on the ubiquitin-ligase domain.
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PMID:Requirements of multiple domains of SLI-1, a Caenorhabditis elegans homologue of c-Cbl, and an inhibitory tyrosine in LET-23 in regulating vulval differentiation. 1107 24

The ubiquitin-protein ligase (E3), hRPF1/Nedd4, is a component of the ubiquitin-proteasome pathway responsible for substrate recognition and specificity. Although previously characterized as a regulator of the stability of cytoplasmic proteins, hRPF1/Nedd4 has also been suggested to have a role in the nucleus. However, in light of the cytoplasmic localization of hRPF1/Nedd4, it is unclear whether bona fide nuclear substrates of hRPF1/Nedd4 exist, and if so, what mechanism may allow a cytoplasmic ubiquitin ligase to manifest nuclear activity. Our search for nuclear substrates led to the identification of the human proline-rich transcript, brain-expressed (hPRTB) protein, the ubiquitination and degradation of which is regulated by hRPF1/Nedd4. Interestingly, hPRTB colocalizes with the splicing factor SC35 in nuclear speckles. Finally, we demonstrate that hRPF1/Nedd4 is indeed capable of entering the nucleus; however, the presence of a functional Rev-like nuclear export sequence in hRPF1/Nedd4 ensures a predominant cytoplasmic localization. Cumulatively, these findings highlight a nuclear role for the ubiquitin ligase hRPF1/Nedd4 and underscore cytoplasmic/nuclear localization as an important regulatory component of hRPF1/Nedd4-substrate recognition.
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PMID:Nuclear import/export of hRPF1/Nedd4 regulates the ubiquitin-dependent degradation of its nuclear substrates. 1134 38

Ubiquitin-mediated protein modification via covalent attachment of ubiquitin has emerged as one of the most common regulatory processes in all eukaryotes. Nedd4-2, closely related to neuronal precursor cell-expressed developmentally down-regulated 4 (Nedd4), is a multimodular ubiquitin-protein ligase comprised of four WW domains and a Hect domain. The WW domains recognize the proline-rich motifs on the multi-subunit amiloride-sensitive epithelial sodium channel (ENaC). To gain insights into the binding of the WW domain to proline-rich peptides, a protein fragment (78 amino acids) containing the fourth WW domain (WW4) of the Nedd4-2 protein was purified and crystallized and X-ray diffraction data were collected. A data set was obtained to 2.5 A resolution from a cryocooled single crystal at a synchrotron source. The crystals belong to the tetragonal space group P4(1)2(1)2 (or P4(3)2(1)2), with unit-cell parameters a = b = 113.43, c = 103.21 A. Analysis of the self-rotation function suggests the presence of four WW4 molecules in the asymmetric unit, with a high unit-cell solvent content of 74%.
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PMID:Crystallization and preliminary X-ray diffraction studies of the WW4 domain of the Nedd4-2 ubiquitin-protein ligase. 1651 Dec 41

A large number of testis-specific genes are involved in the complex process of mammalian spermatogenesis. Identification of these genes and their roles is important for understanding the mechanisms underlying spermatogenesis. Here we report on a novel human RING finger protein, ZNF645, which contains a C3HC4 RING finger domain, a C2H2 zinc-finger domain, and a proline-rich region, indicating that it has a structure similar to that of the c-Cbl-like protein Hakai. ZNF645 was exclusively expressed in normal human testicular tissue. Immunohistochemical analysis confirmed that ZNF645 protein was present in spermatocytes, round and elongated spermatids, and Leydig cells. Immunofluorescence staining of mature sperms further showed that the ZNF645 protein was localized over the postacrosomal perinuclear theca region and the entire length of sperm tail. An in vitro ubiquitination assay indicated that the RING finger domain of the ZNF645 protein had E3 ubiquitin ligase activity. Therefore, we suggest that ZNF645 might act as an E3 ubiquitin-protein ligase and play a role in human sperm production and quality control.
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PMID:Human RING finger protein ZNF645 is a novel testis-specific E3 ubiquitin ligase. 2065 3