Basidiomycete Lentinula edodes CDC5 and a novel interacting protein CIPB bind to a newly isolated target gene in an unusual manner

The homologue of Lentinula edodes ctg1, a target for CDC5 and its interacting partner CIPB, from Coprinopsis cinerea is involved in fruitingbody morphogenesis of C. cinerea

“…The Cc.cdc5 cDNA fragment (nt 1701-2469) was inserted into pET-HA and the resulting plasmid was introduced into E. coli BL21 DE3 (plysS) (Novagen), producing HA-Cc.Cdc5 (aa 568-822) as a prey protein. The GST-pulldown assay was performed according to the procedure reported previously (Nakazawa et al 2008a). The pulled-down HA-Cc.Cdc5 (568-822) was detected by immunoblotting using anti-HA-antibody F-7 (Santa Cruz Biotechnology, Santa Cruz, CA, USA).…”

“…To investigate the molecular mechanism underlying the fruiting development of the basidiomycetous mushroom Lentinula edodes (Berk.) Pegler, commonly called "shiitake," a variety of genes (or cDNAs) specifi c for fruiting development have been isolated from L. edodes (Leung et al 2000;Miyazaki et al 2005;Chum et al 2008), and their properties and functions have been analyzed (Lee et al 2007;Nakazawa et al 2008a). One of these, the Le.cdc5 gene (Miyazaki et al 2004), is a homologue of the Schizosaccharomyces pombe cdc5 + gene (Ohi et al 1994).…”

“…To demonstrate that the Le.CDC5 protein functions as a transcription factor, we previously attempted to isolate its target gene(s). A genomic binding site (GBS) cloning experiment (Inoue et al 1991) with the Le.CDC5 (1-185) peptide and an inverse polymerase chain reaction (PCR) resulted in the isolation of a target gene designated ctg1 (CDC5 target gene) (Nakazawa et al 2008a). It contains a 7-bp consensus-like sequence (5′-GCAATCT-3′) for Le.CDC5 binding in its transcribed region downstream of the start codon.…”

“…At least two each of the upstream and downstream sequences appear to bind CIPB in vitro. These fi ndings suggest that Le.CDC5 and CIPB can cooperatively regulate the expression of ctg1 (Nakazawa et al 2008a). The ctg1 gene product (CTG1; 159 aa residues) contains a short leucine zipper-like motif, LALX 5 LX 6 L, at the N-terminus and a short basic amino acid sequence, KKK, near the C-terminus.…”

“…The ctg1 gene product (CTG1; 159 aa residues) contains a short leucine zipper-like motif, LALX 5 LX 6 L, at the N-terminus and a short basic amino acid sequence, KKK, near the C-terminus. Northern blot analysis suggested that the ctg1 gene is transcribed most actively in the upper halves of the stipes of fruiting bodies, implying its involvement in regulation of stipe elongation (Nakazawa et al 2008a).…”