Ataxia-telangiectasia mutated (ATM) is a cell cycle checkpoint kinase that upon activation by DNA damage leads to cell cycle arrest and DNA repair or apoptosis. The absence of Atm or the occurrence of loss-of-function mutations in Atm predisposes to tumorigenesis. MAPK7 has been implicated in numerous types of cancer with pro-survival and pro-growth roles in tumor cells, but its functional relation with tumor suppressors is not clear. In this study, we show that absence of MAPK7 delays death due to spontaneous tumor development in Atm-/-mice. Compared with Atm-/-thymocytes, Mapk7-/

SUMMARY The cellular response to DNA double-strand breaks (DSBs) is mobilized by the protein kinase ATM, which phosphorylates key players in the DNA damage response (DDR) network. A major question is how ATM controls DSB repair. Optimal repair requires chromatin relaxation at damaged sites. Chromatin reorganization is coupled to dynamic alterations in histone posttranslational modifications. Here, we show that in human cells, DSBs induce monoubiquitylation of histone H2B, a modification that is associated in undamaged cells with transcription elongation. We find that this process relies on recruitment to DSB sites and ATM-dependent phosphorylation of the responsible E3 ubiquitin ligase: the RNF20-RNF40 heterodimer. H2B monoubiquitylation is required for timely recruitment of players in the two major DSB repair pathways—nonhomologous end-joining and homologous recombination repair—and optimal repair via both pathways. Our data and previous data suggest a two-stage model for chromatin decondensation that facilitates DSB repair.

MUC1, a glycoprotein overexpressed by a variety of human adenocarcinomas, is a type I transmembrane protein (MUC1/TM) that soon after its synthesis undergoes proteolytic cleavage in its extracellular domain. This cleavage generates two subunits, ␣ and ␤, that specifically recognize each other and bind together in a strong noncovalent interaction. Proteolysis occurs within the SEA module, a 120-amino acid domain that is highly conserved in a number of heavily glycosylated mucin-like proteins. Post-translational cleavage of the SEA module occurs at a site similar to that in MUC1 in the glycoproteins IgHepta and MUC3. However, as in the case of other proteins containing the cleaved SEA module, the mechanism of MUC1 proteolysis has not been elucidated. Alternative splicing generates two transmembrane MUC1 isoforms, designated MUC1/Y and MUC1/X. We demonstrated here that MUC1/X, whose extracellular domain is comprised solely of the SEA module in addition to 30 MUC1 N-terminal amino acids, undergoes proteolytic cleavage at the same site as the MUC1/TM protein. In contrast, the MUC1/Y isoform, composed of an N-terminally truncated SEA module, is not cleaved. Cysteine or threonine mutations of the MUC1/X serine residue (Ser-63) immediately C-terminal to the cleavage site generated cleaved proteins, whereas mutation of the Ser-63 residue of MUC1/X to any other of 17 amino acids did not result in cleavage. In vitro incubation of highly purified precursor MUC1/X protein resulted in self-cleavage. Furthermore, addition of hydroxylamine, a strong nucleophile, markedly enhanced cleavage. Both these features are signature characteristics of self-cleaving proteins, and we concluded that MUC1 undergoes autoproteolysis mediated by an N 3 O-acyl rearrangement at the cleavage site followed by hydrolytic resolution of the unstable ester and concomitant cleavage. It is likely that all cleaved SEA module-containing proteins follow a similar route.The MUC1 gene is highly expressed in a number of human epithelial malignancies, including breast, prostate, and colon carcinomas, as well as on the malignant plasma cells of multiple myeloma (1-9). As a well characterized tumor-associated protein, it has generated considerable interest as a tumor marker for disease prognosis (10 -14) as well as a target for tumor cell killing (15-18). Although alternative splicing can generate multiple MUC1 protein forms (19 -23), the most intensively studied MUC1 protein is a type I transmembrane protein comprised of a heavily glycosylated extracellular domain containing a tandem-repeat array, a transmembrane domain, and a cytoplasmic domain (Fig. 1, MUC1/TM) (24 -26). MUC1/TM is proteolytically cleaved soon after its synthesis, generating two subunits, ␣ and ␤, that specifically recognize each other and bind together by a strong noncovalent interaction (27).Cleavage occurs within the SEA module (28 -30), a highly conserved protein module so-called from its initial identification in a sperm protein, in enterokinase, and in agrin (31), that is found in a numb...

The isolation and characterization of complementary DNAs (cDNAs) which code for an epithelial antigen aberrantly expressed in human breast tumor tissue are described here. The only information regarding the primary structure of this potentially important antigen has been a 20-amino-acid repeat motif. We now report the complete amino acid sequences of different forms of the human epithelial tumor antigen as deduced from the nucleotide sequence of isolated non-repeat cDNAs. The diversity of protein forms is generated by a series of alternative splicing events that occur in the regions located upstream and downstream to a central tandem repeat array. Isolated cDNAs coding for the upstream region show that differential usage of alternative splice acceptor sites may generate two protein forms containing putative signal peptides of varying hydrophobicities. The complexity of possible antigen forms is further compounded by alternative splicing events occurring in the region 3' to the repeat array. The isolated cDNAs 3' to the tandem repeats indicate that whereas one mRNA transcript is colinear with the gene, and defines an open reading frame (ORF) containing 160 amino acids downstream to the repeat array, a second cDNA correlates with a mRNA that is generated by a series of splicing events. The deduced amino acid sequence of the spliced cDNA contains an ORF that is identical for 149 amino acids downstream to the repeat array with the amino acid sequence of the unspliced cDNA. At this point it diverges and continues for an additional 179 amino acids. The sequence contains a highly hydrophobic 28-amino-acid peptide, located towards the carboxyl terminus, that may correspond to a transmembrane region. The cDNAs and deduced amino acid sequences, presented here, define the complete amino acid sequences of the epithelial tumor antigen and demonstrate the existence of multiple protein forms that probably localize to different cellular and extracellular compartments.