53BP1 is a Positive Regulator of the BRCA1 Promoter

Germline mutations in the BRCA1 tumor suppressor gene contribute to familial breast and ovarian tumor formation. Sporadic breast and ovarian cancer, however, which accounts for more than 90% of total cases and virtually lacks BRCA1 mutations, exhibits reduced expression of the BRCA1 gene. The magnitude of this reduction correlates with disease progression. In this report we have identified an imperfect palindrome sequence for binding of the 53BP1-containing complex, -40TTCCGTGG CAACGGAA-25, within the BRCA1 minimal promoter. Overexpression of 53BP1 activates a luciferase reporter driven by the wild type BRCA1 minimal promoter, but not by the BRCA1 minimal promoter with mutated palindrome sequence. Depletion of endogenous 53BP1 by siRNA suppresses activity of the BRCA1 minimal promoter. In vitro and in vivo DNA-protein interaction studies demonstrate that this palindrome sequence binds to the 53BP1-containing complex. These findings establish a positive regulation of the BRCA1 promoter by 53BP1. Disruption of this regulation in cancer cells may provide a molecular mechanistic basis for sporadic breast and ovarian tumor formation.     

Fibrillin-1 regulates the bioavailability of TGFbeta1

We have discovered that fibrillin-1, which forms extracellular microfibrils, can regulate the bioavailability of transforming growth factor (TGF) beta1, a powerful cytokine that modulates cell survival and phenotype. Altered TGFbeta signaling is a major contributor to the pathology of Marfan syndrome (MFS) and related diseases. In the presence of cell layer extracellular matrix, a fibrillin-1 sequence encoded by exons 44-49 releases endogenous TGFbeta1, thereby stimulating TGFbeta receptor-mediated Smad2 signaling. This altered TGFbeta1 bioavailability does not require intact cells, proteolysis, or the altered expression of TGFbeta1 or its receptors. Mass spectrometry revealed that a fibrillin-1 fragment containing the TGFbeta1-releasing sequence specifically associates with full-length fibrillin-1 in cell layers. Solid-phase and BIAcore binding studies showed that this fragment interacts strongly and specifically with N-terminal fibrillin-1, thereby inhibiting the association of C-terminal latent TGFbeta-binding protein 1 (a component of the large latent complex [LLC]) with N-terminal fibrillin-1. By releasing LLC from microfibrils, the fibrillin-1 sequence encoded by exons 44-49 can contribute to MFS and related diseases.     

PAK1 negatively regulates the activity of the Rho exchange factor NET1

Rho family small G-protein activity is controlled by guanine nucleotide exchange factors that stimulate the release of GDP, thus allowing GTP binding. Once activated, Rho proteins control cell signaling through interactions with downstream effector proteins, leading to changes in cytoskeletal organization and gene expression. The ability of Rho family members to modulate the activity of other Rho proteins is also intrinsic to these processes. In this work we show that the Rac/Cdc42hs-regulated protein kinase PAK1 down-regulates the activity of the RhoA-specific guanine nucleotide exchange factor NET1. Specifically, PAK1 phosphorylates NET1 on three sites in vitro: serines 152, 153, and 538. Replacement of serines 152 and 153 with glutamate residues down-regulates the activity of NET1 as an exchange factor in vitro and its ability to stimulate actin stress fiber formation in cells. Using a phospho-specific antibody that recognizes NET1 phosphorylated on serine 152, we show that PAK1 phosphorylates NET1 on this site in cells and that Rac1 stimulates serine 152 phosphorylation in a PAK1-dependent manner. Furthermore, coexpression of constitutively active PAK1 inhibits the ability of NET1 to stimulate actin polymerization only when serines 152 and 153 are present. These data provide a novel mechanism for the control of RhoA activity by Rac1 through the PAK-dependent phosphorylation of NET1 to reduce its activity as a guanine nucleotide exchange factor.     

Skp1-Cullin-F-box-dependent degradation of Aah1p requires its interaction with the F-box protein Saf1p

When yeast cells enter into quiescence in response to nutrient limitation, the adenine deaminase Aah1p is specifically degraded via a process requiring the F-box protein Saf1p and components of the Skp1-Cullin-F-box complex. In this paper, we show that Saf1p interacts with both Aah1p and Skp1p. Interaction with Skp1p, but not with Aah1p, requires the F-box domain of Saf1p. Based on deletion and point mutations, we further demonstrate that the F-box domain of Saf1p is critical for degradation of Aah1p. We also establish that overexpression of Saf1p in proliferating cells is sufficient to trigger the degradation of Aah1p. Using this property and a two-dimensional protein gel approach, we found that Saf1p has a small number of direct targets. Finally, we isolated and characterized several point mutations in Aah1p, which increase its stability during quiescence. The majority of the mutated residues are located in two distinct exposed regions in the Aah1p three-dimensional model structure. Two hybrid experiments strongly suggest that these domains are directly involved in interaction with Saf1p. Importantly, we obtained a mutation in Aah1p that does not affect the protein interaction with Saf1p but abolishes Aah1p degradation. Because this mutated residue is an exposed lysine in the Aah1p three-dimensional model, we propose that it is likely to be a major ubiquitylation site. All together, our data strongly argue for Saf1p being a bona fide Skp1-Cullin-F-box subunit.     

单核细胞趋化蛋白诱导蛋白-1的研究进展

单核细胞趋化蛋白诱导蛋白-1（MCPIP1）是最近发现的一类具有免疫调节作用的CCCH型锌指家族分子。MCPIP1可被LPS、IL-1β或MCP-1等多种炎性因子刺激表达,可通过下调炎症因子（如IL-6、IL-12p40等）表达,从而负向调控炎症过程。MCPIP1的作用机制主要是作为RNA酶调节某些炎性因子mRNA或pre-miRNA的降解。此外,MCPIP1也可作为去泛素化酶靶向TNF受体相关蛋白（TRAFs）成员,从而负向调控JNK和NF-κB信号活化。将从MCPIP1分子的发现、基因和蛋白质结构、生物学功能、表达调控以及临床应用前景等几个方面进行阐述。     

NF1-RAC1 axis regulates migration of the melanocytic lineage

Metastases's spreading is the main cause of mortality for advanced stage cancer patients, including melanoma. The formation of metastases is favored by enhanced migratory and invasive capacities of tumor cells. Tumor suppressor gene NF1 is a negative regulator of RAS and its deregulation plays an important role in several aspects of melanoma transformation and progression. However, very little is described about the role of NF1 in cellular migration and invasion. In this study, our results show on the one hand, that the loss of NF1 expression delays migration of human melanoblasts via a RAC1-dependent mechanism. On the other hand, our data indicate that NF1 loss in melanoma cells is enhancing migration, intravasation and metastases formation in vivo. Moreover, not only this phenotype is associated with an upregulation of PREX1 but also patient-derived melanoma samples with low NF1 expression present increased levels of PREX1. In sum, our study brings new elements on the mechanism controlling cellular migration in the context of NF1 loss. These data are of prime interest to improve treatment strategies against all NF1-mutated tumors, including this subtype of melanoma.     

AXR1-ECR1-dependent conjugation of RUB1 to the Arabidopsis Cullin AtCUL1 is required for auxin response

Mutations in the AXR1 gene result in a reduction in auxin response and diverse defects in auxin-regulated growth and development. In a previous study, we showed that AXR1 forms a heterodimer with the ECR1 protein. This enzyme activates the ubiquitin-related protein RUB1 in vitro. Furthermore, we showed that the Skp1-Cul1/Cdc53-F-box (SCF) subunit AtCUL1 is modified by RUB1 in vivo. In this report, we demonstrate that the formation of RUB-AtCUL1 is dependent on AXR1 and ECR1 in vivo. The expression of AXR1 and ECR1 is restricted to zones of active cell division and cell elongation, consistent with their role in growth regulation. These results provide strong support for a model in which RUB conjugation of AtCUL1 affects the function of SCF E3s that are required for auxin response.     

Structure and localization of the human SULT1B1 gene: neighborhood to SULT1E1 and a SULT1D pseudogene.

The soluble sulfotransferases are involved in the elimination of xenobiotics, the activation of procarcinogens, and the regulation of hormones. They comprise a gene superfamily (SULT). The structure and chromosomal location of nine human SULT genes are known. We have characterized a further gene, SULT1B1. Its structure is similar to that of other SULT1 genes. However, the total length of its eight exons and the introns (33.6 kb) is larger than that of other human SULT1 genes (4 to 21 kb). The SULT1B1 gene sequence is part of a sequence entry in the unfinished High-Throughput Genomic Sequences (HTGS) division of GenBank. However, the order and orientation of the SULT1B1 exons are not correct in this entry. SULT1B1 is located on chromosome 4q13.1, nearly 100 kb downstream of SULT1E1 on the same strand. The intervening sequence contains a SULT-like structure showing substantial homology to the mouse SULT1D1 cDNA recently described. However, in humans this structure represents a pseudogene (SULT1D1P) because of mutated splice donors/acceptors and in-frame stop codons in the sequence corresponding to exon II. This SULT gene cluster is located on the minus strand of chromosome 4 with SULT1B1 being closest to the centromer.