Isolation and characterization of cDNA clones encoding polypeptides related to a Dictyostelium discoideum cyclic AMP binding protein

Abstract

By screening a cDNA library with a cDNA encoding the Dicfyostefium discoideum CAMP-binding protein CABP1, under conditions of reduced stringency, we have isolated clones which code for two closely related molecules. Hybrid selection experiments indicated that these cDNAs encoded polypeptides with molecular masses of 34 (p34) and 31 (p31) kDa, both of which were recognized by anti-CABP1 monoclonal antibodies. Sequence analysis revealed that the clones were identical except for the presence of a 102 nucleotide segment inserted in-frame in the p34 cDNAs, just downstream of the translation initiation codon. DNA blot analysis suggested that p34 and p31 were encoded by the same gene. This hypothesis was strongly supported by the observation that both polypeptides were generated when a single cDNA was expressed under the control of the actin 15 promoter in D. discoideum cells. RNA blot analysis indicated that the cDNAs were complementary to three developmentally regulated transcripts of sizes 1.15 kb, 1-25 kb and 1.4 kb. Comparison of the derived amino acid sequences of p34 and p31 with those of the two subunits of CABPl indicated that these polypeptides were very closely related, and that the corresponding genes probably arose by duplication followed by sequence divergence. Finally, the carboxy termini of these four polypeptides demonstrated 50% similarity to two polypeptides encoded by a bacterial plasmid which confers resistance to tellurium anions.

Spliceosomal genes in the D. discoideum genome: a comparison with those in H. sapiens, D. melanogaster, A. thaliana and S. cerevisiae

“…Our sequence analyses of genomic and EST databases strongly support earlier findings (Grant and Tsang, 1990; Bain et al, 1991; Greenwood and Tsang, 1991; Escalante et al, 2003) that D. discoideum has bona fide alternative splicing. To date, we have examined nearly all 13,527 genes individually and compared them with the available EST and cDNAs.…”

Dictyostelium TRFA Homologous to Yeast Ssn6 Is Required for Normal Growth and Early Development

“…Southern and Northern Blot Analyses-Genomic DNA was isolated from Dictyostelium cells as described (28). Digested DNA was separated by 0.6% agarose gel and then transferred to a Fine Trap NT-21 membrane (Nihon Eido Co., Japan) by electroblotting.…”