Antisense oligonucleotide inhibition of acetylcholinesterase gene expression induces progenitor cell expansion and suppresses hematopoietic apoptosis ex vivo.
To examine the role of acetylcholinesterase (EC 3.1.1.7) in hematopoietic cell proliferation and differentiation, we administered a 15-mer phosphorothioate oligonucleotide, antisense to the corresponding ACHE gene (AS-ACHE), to primary mouse bone marrow cultures. Within 2 hr of AS-ACHE addition to the culture, ACHE mRNA levels dropped by approximately 90%, as compared with those in cells treated with the "sense" oligomer, S-ACHE. Four days after AS-ACHE treatment, ACHE mRNA increased to levels 10-fold higher than in S-ACHE cultures or in fresh bone marrow. At this later time point, differential PCR display revealed significant differences between cellular mRNA transcripts in bone marrow and those in AS-ACHE- or S-ACHE-treated cultures. These oligonucleotide-triggered effects underlay considerable alterations at the cellular level: AS-ACHE but not S-ACHE increased cell counts, reflecting enhanced proliferation. In the presence of erythropoietin it also enhanced colony counts, reflecting expansion of progenitors. AS-ACHE further suppressed apoptosis-related fragmentation of cellular DNA in the progeny cells, and it diverted hematopoiesis toward production of primitive blasts and macrophages in a dose-dependent manner promoted by erythropoietin. These findings suggest that the hematopoietic role of acetylcholinesterase, anticipated to be inverse to the observed antisense effects, is to reduce proliferation of the multipotent stem cells committed to erythropoiesis and megakaryocytopoiesis and macrophage production and to promote apoptosis in their progeny. Moreover, these findings may explain the tumorigenic association of perturbations in ACHE gene expression with leukemia.
Soreq
H
H
Department of Biological Chemistry, Hebrew University of Jerusalem, Israel.
Patinkin
D
D
Lev-Lehman
E
E
Grifman
M
M
Ginzberg
D
D
Eckstein
F
F
Zakut
H
H
D000110:Acetylcholinesterase / Q000096:biosynthesis* / Q000235:genetics; D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D001483:Base Sequence; D001854:Bone Marrow Cells; D002454:Cell Differentiation / Q000187:drug effects; D002478:Cells, Cultured; D003114:Colony-Forming Units Assay; D004268:DNA-Binding Proteins / Q000096:biosynthesis; D004790:Enzyme Induction; D050981:Erythroid-Specific DNA-Binding Factors; D004921:Erythropoietin / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects*; D006412:Hematopoietic Stem Cells / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology; D007377:Interleukin-3 / Q000494:pharmacology; D007700:Kinetics; D008264:Macrophages / Q000166:cytology / Q000187:drug effects; D051379:Mice; D008969:Molecular Sequence Data; D016376:Oligonucleotides, Antisense / Q000494:pharmacology*; D012333:RNA, Messenger / Q000096:biosynthesis; D014157:Transcription Factors / Q000096:biosynthesis
D004268:DNA-Binding Proteins; D050981:Erythroid-Specific DNA-Binding Factors; D007377:Interleukin-3; D016376:Oligonucleotides, Antisense; D012333:RNA, Messenger; D014157:Transcription Factors; D004921:Erythropoietin; D000110:Acetylcholinesterase