Targeted disruption of H2B-V encoding a particular H2B histone variant causes changes in protein patterns on two-dimensional polyacrylamide gel electrophoresis in the DT40 chicken B cell line.
The chicken H2B gene family comprises eight members (H2B-I to H2B-VIII), which are all located in two major histone gene clusters. All of them have been shown to encode four different protein variants (classes I to IV). In the DT40 chicken B cell line, the H2B-V gene, encoding the class III H2B variant, constituted about 10% of the total intracellular mRNA from all the H2B genes. To study the nature of this particular variant in vivo, we generated heterozygous (H2B-V, +/-) and homozygous (H2B-V, -/-) DT40 mutants by targeted integration. The remaining H2B genes were shown to be expressed more in these mutants than in the wild-type cell lines. The growth rate of DT40 cells was unchanged in the absence of the H2B-V gene. Two-dimensional polyacrylamide gel electrophoresis showed that the protein patterns were, on the whole, similar between the wild-type and homozygous cell lines. However, within this constant background, some cellular proteins disappeared or decreased quantitatively in the homozygous mutants, and several other proteins increased or newly appeared. These results suggest that the class III H2B variant participates negatively or positively in regulation of the expression of particular genes that encode the proteins that vary in DT40 cells. This type of regulation is possibly mediated through alterations in nucleosome structure over the restricted regions involving the putative genes of the DT40 genome.
Department of Biochemistry, Miyazaki Medical College, Japan.
Y
Y
Takami
S
S
Takeda
T
T
Nakayama
D000818:Animals; D001402:B-Lymphocytes; D002460:Cell Line; D002645:Chickens; D015180:Electrophoresis, Gel, Two-Dimensional; D004591:Electrophoresis, Polyacrylamide Gel; D015870:Gene Expression; D014644:Genetic Variation; D006657:Histones / Q000096:biosynthesis* / Q000235:genetics* / Q000302:isolation & purification; D005810:Multigene Family; D012333:RNA, Messenger / Q000032:analysis / Q000096:biosynthesis; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000302:isolation & purification; D015183:Restriction Mapping; D014162:Transfection
D006657:Histones; D012333:RNA, Messenger; D011994:Recombinant Proteins