CHARACTERIZATION OF THE DICTYOSTELIUM-DISCOIDEUM CELLULOSE-BINDING PROTEIN CELB AND REGULATION OF GENE-EXPRESSION

Similar to other stages of Dictyostelium development, spore germinatio n is a particularly suitable model for studying regulation of gene exp ression, The transition from spore to amoeba is accompanied by develop mentally regulated changes in both protein and mRNA synthesis, A numbe r of spore germination-specific cDNAs have been isolated previously, A mong these are two members of the 270 gene family, a group of four gen es defined by the presence of a common tetrapeptide repeat of Thr-Glu- Thr-Pro, celA (formerly called 270-6) and celB (formerly 270-11) are e xpressed solely and coordinately during spore germination, the levels of the respective mRNAs being low in dormant spores, rising during gem ination to a maximum level at about 2 h, and then rapidly declining as amoebae are released from spores, The mRNAs are not found ill growing cells or during multicellular development, The rapidity with which th ese transcripts accumulate and then disappear during germination impli es that the respective products may be important for the process, We r eported previously that the CelA protein is a cellulase (endo-1,4-beta -glucanase (EC 3.2.1.4)). In the present investigation, properties of the CelB protein, a glycosylated protein of 532 amino acids, 36% of wh ich are serine or threonine, were examined, and the upstream sequences involved in the developmental regulation of the expression of the gen e have been determined, The CelB protein does not demonstrate cellulas e activity, but it has a cellulose-binding domain, Its role, if any, i n degradation of tile cellulose-containing spore wall is unknown. To i dentify cis-acting elements in the celB promoter, unidirectional 5' de letions of the celB upstream noncoding region were constructed and use d to transform amoebae, Analysis of promoter activity during different stages of development shows that a short, very APT-rich sequence of a pproximately 81 base pairs is sufficient for spore-specific celB trans cription, Contained in this sequence is the Myb oncogene protein bindi ng site, TAACTG, which was shown previously to be a negative regulator of celA transcription, Dictyostelium and mouse Myb proteins bind to t his region of the promoter, suggesting that Myb might regulate celB ge ne expression negatively as it does in celA.