H2A.X. a histone isoprotein with a conserved C-terminal sequence, is encoded by a novel mRNA with both DNA replication type and polyA 3′ processing signals

Abstract

A full length cDNA clone that directs the in vitro synthesis of human histone H2A isoprotein H2A.X has been isolated and sequenced. H2A.X contains 142 amino acid residues, 13 more than human H2A.1. The sequence of the first 120 residues of H2A.X is almost identical to that of human H2A.1. The sequence of the carboxy-terminal 22 residues of H2A.X is unrelated to any known sequence in vertebrate histone H2A; however, it contains a sequence homologous with those of several lower eukaryotes. This homology centers on the carboxy-terminal tetrapeptide which in H2A.X is SerGlnGluTyr. Homologous sequences are found in H2As of three types of yeasts, in Tetrahymena and Drosophila. Seven of the nine carboxy-terminal amino acids of H2A.X are identical with those of S. cerevisiae H2A.1. It is suggested that this H2A carboxy-terminal motif may be present in all eukaryotes. The H2A.X cDNA is 1585 bases long followed by a polyA tail. There are 73 nucleotides in the 5' UTR, 432 in the coding region, and 1080 in the 3' UTR. Even though H2A.X is considered a basal histone, being synthesized in G1 as well as in S-phase, and its mRNA contains polyA addition motifs and a polyA tail, its mRNA also contains the conserved stem-loop and U7 binding sequences involved in the processing and stability of replication type histone mRNAs. Two forms of H2A.X mRNA, consistent with the two sets of processing signals were found in proliferating cell cultures. One, about 1600 bases long, contains polyA; the other, about 575 bases long, lacks polyA. The short form behaves as a replication type histone mRNA, decreasing in amount when cell cultures are incubated with inhibitors of DNA synthesis, while the longer behaves as a basal type histone mRNA.

H2AX: the histone guardian of the genome

“…First identified in 1980 in human cells as an electrophoretic isoform of the core histone H2A [2], H2AX was subsequently sequenced in the late 80's [3]. These early studies noted the presence of a short COOH terminal tail not found in any other mammalian H2A isoforms, but being present-with various degrees of homology and linker lengths-in the unique H2A homologue of lower eukaryotes [3,4].…”

“…These early studies noted the presence of a short COOH terminal tail not found in any other mammalian H2A isoforms, but being present-with various degrees of homology and linker lengths-in the unique H2A homologue of lower eukaryotes [3,4]. H2AX constitutes a major H2A species, and its levels vary from 2-25% of the mammalian histone H2A pool depending on the cell line or tissue examined [5,6].…”

Low-dose radiation: Thresholds, bystander effects, and adaptive responses

“…More-sensitive assays are being developed, an important one being the ability to detect the cellular presence of the most serious and potentially lethal type of cellular damage, the DNA doublestrand break (DSB). This assay is based on the finding that one of the highly conserved histone proteins that package the DNA into chromatin, H2AX, becomes phosphorylated at the sites of nascent DNA DSBs (5)(6)(7)(8). The response is highly amplified and rapid, involving the phosphorylation of hundreds to thousands of H2AX molecules within minutes on several megabase equivalents of chromatin flanking the DSB.…”

Two Types of Polyadenated mRNAs are Synthesized from Drosophila Replication‐Dependent Histone Genes

“…In some cases, such mRNAs represent a very minor part of the total mRNA pool (Levine et al, 1987). In several histone H I transcripts in chicken (Kirsh et al, 1989) and mouse (Cheng et al, 1989), as well as in transcripts encoding the histone H2A.X variant (Mannironi et al, 1989;Nagata et al, 1991), the two modes of the 3' processing occur with comparable efficiency in the same cell or tissue type. In all these cases the polyadenylated messengers behave as replacement type histone mRNAs, which are not destabilized at the end of the S-phase, in contrast to the nonpolyadenylated mRNAs.…”

“…In all these cases the polyadenylated messengers behave as replacement type histone mRNAs, which are not destabilized at the end of the S-phase, in contrast to the nonpolyadenylated mRNAs. The existence of such poly(A)-containing mRNAs is probably necessary to ensure the basal synthesis of the respective histone variants (Kirsh et al, 1989;Cheng et al, 1989;Mannironi et al, 1989;Nagata et al, 1991).…”