Involvement of a short interspersed element in epigenetic transcriptional silencing of the amoebapore gene in Entamoeba histolytica.
Transcriptional silencing of an amoebapore (ap-a) gene occurred in Entamoeba histolytica following the transfection of plasmids containing a DNA segment (473 bp) homologous to the 5' upstream region of the gene (R. Bracha, Y. Nuchamowitz, and D. Mirelman, Eukaryot. Cell 2:295-305, 2003). This segment contains the promoter region of the ap-a gene, a T-rich stretch, followed by a truncated SINE1 (short interspersed element 1) that is transcribed from the antisense strand. Transfection of plasmids containing truncated SINE1 sequences which lack their 3' regulatory elements upstream of the ap-a gene was essential for the downstream silencing of the ap-a gene while transfection with plasmids containing the entire SINE1 sequence or without the T-rich stretch promoted the overexpression of the ap-a gene. Both the T-rich stretch and sequences of the 5' SINE1 were essential for the transcription of SINE1. RNA extracts from gene-silenced cultures showed small amounts of short (approximately 140-nucleotide), single-stranded molecules with homology to SINE1 but no short interfering RNA. Chromatin immunoprecipitation analysis with an antibody against methylated K4 of histone H3 showed a demethylation of K4 at the domain of the ap-a gene, indicating transcriptional inactivation. These results suggest the involvement of SINE1 in triggering the gene silencing and the role of histone modification in its epigenetic maintenance.
Anbar
Michael
M
Department of Biological Chemistry, Weizmann Institute of Science, P.O. Box 26, Rehovot 76100, Israel.
Bracha
Rivka
R
Nuchamowitz
Yael
Y
Li
Yan
Y
Florentin
Anat
A
Mirelman
David
D
D000818:Animals; D003596:Cytosine / Q000378:metabolism; D004748:Entamoeba histolytica / Q000235:genetics* / Q000502:physiology; D044127:Epigenesis, Genetic; D020868:Gene Silencing; D017930:Genes, Reporter; D006657:Histones / Q000378:metabolism; D007473:Ion Channels / Q000235:genetics* / Q000378:metabolism; D016366:Open Reading Frames; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D015800:Protozoan Proteins / Q000235:genetics* / Q000378:metabolism; D012313:RNA / Q000235:genetics / Q000378:metabolism; D020086:Short Interspersed Nucleotide Elements; D014158:Transcription, Genetic
D006657:Histones; D007473:Ion Channels; D015800:Protozoan Proteins; C041670:amoebapore proteins, protozoan; D012313:RNA; D003596:Cytosine