Irreversible Repression of DNA Synthesis in Fanconi Anemia Cells Is Alleviated by the Product of a Novel Cyclin-Related Gene

Abstract

Primary fibroblasts from patients with the genetic disease Fanconi anemia, which are hypersensitive to cross-linking agents, were used to screen a cDNA library for sequences involved in their abnormal cellular response to a cross-linking challenge. By using library partition and microinjection of in vitro-transcribed RNA, a cDNA clone, pSPHAR (S-phase response), which is able to correct the permanent repression of semiconservative DNA synthesis rates characteristic of these cells, was isolated. Wild-type SPHAR mRNA is expressed in all fibroblasts so far analyzed, including those of Fanconi anemia patients. Correction of the abnormal response in these cells appears therefore to be due to overexpression after cDNA transfer rather than to genetic complementation. The cDNA contains an open reading frame coding for a polypeptide of 7.5 kDa. Rabbit antiserum directed against a SPHAR peptide detects a protein of 7.9 kDa in Western blots (immunoblots) of whole-cell extracts from proliferating, but not resting, fibroblasts. The deduced amino acid sequence of SPHAR contains a motif found in the cyclins, and it is proposed that SPHAR acts within the injected cell by interfering with the cyclin-controlled maintenance of S phase. In agreement with this proposal, normal cells transfected with an antisense SPHAR expression vector have a significantly reduced rate of DNA synthesis during S phase and a prolonged G 2 phase, reflecting the need for postreplicative DNA processing before entry into mitosis.

Distinctive gene expression profiles of CD34 cells from patients with myelodysplastic syndrome characterized by specific chromosomal abnormalities

“…37,75 Other genes prominently dysregulated in monosomy 7 cells promote cell proliferation and differentiation, including the cell cycle regulator SPHAR, the DNA replication checkpoint gene Rad17, and the signal transduction gene TPO; in contrast, genes with broadly antagonistic functions were down-regulated, including the cyclin-dependent kinase inhibitor p21, several signal transducers such as GATA2, and members of the mammalian mitogen-activated protein (MAP) family. [80][81][82] In general, the gene expression pattern in monosomy 7 progenitor cells was consistent with functional characteristics of high proliferation and malignant potential.…”

Mechanistic insight into taxol-induced cell death

“…Loss of SPHAR function reduces the rate of DNA synthesis during S phase and causes G2-phase prolongation (Digweed et al, 1995). NAP-1 is important for chromatin assembly and cell cycle regulation.…”

Cloning of two candidate tumor suppressor genes within a 10 kb region on chromosome 13q14, frequently deleted in chronic lymphocytic leukemia

“…The signal element for mRNA polyadenylation is underlined 13q candidate tumor suppressor genes Y Liu et al 10 kD respectively. Similarly sized proteins have previously been shown to be involved in malignant transformation and growth control (Soulier et al, 1994;Fu et al, 1994;Digweed et al, 1995;Kishida et al, 1995). It can of course not be completely ruled out that the short ORFs may be due to incomplete transcripts.…”