Differential gene expression in Physarum.

Physarum polycephalum has a life-cycle that encompasses two very distinct vegetatively growing cell-types; uninucleate microscopic amoebae, and multinucleate macroscopic plasmodia. cDNA libraries have been prepared from both amoebae and plasmodia using the M13 vector mp8. Clones from the two libraries were screened using a differential hybridisation procedure that identifies clones derived from mRNAs much more abundant in one cell-type than in the other. For both the amoebal library and the plasmodial library it was found that about 5% of the clones represented genes preferentially expressed in the cell-type from which the library was prepared. Some of the cell-type-specific clones obtained were used to probe northern blots of amoebal and plasmodial RNA. Two of the plasmodial-specific clones were found to be derived from highly abundant mRNAs, constituting between 0.5% and 2% of total plasmodial mRNA. Selected clones were then used to look at changes in mRNA concentrations during development by probing northern blots of RNA from amoebae, plasmodia and intermediate cell-types. It was found that the plasmodial-specific mRNAs examined fell into two classes; those expressed early in development, and those expressed late in development. The amoebal-specific clones analysed constituted a single group, with each of the probes used detecting an mRNA whose concentration declined markedly in early development. Some of the changes in gene expression were examined more quantitatively by dot blotting. It was found that the difference in concentration of cell-type-specific mRNAs between amoebae and plasmodia varied from between 10 fold to greater than 100 fold. Analysis of the pattern of gene expression was begun in two mutant strains of Physarum which are unable to complete development. Results obtained from a strain which is arrested late in development (RA612) suggest that the developmentally arrested cells express the plasmodial-specific genes which are activated early in development, but not those that become active late in development. A few of the clones have been sequenced, but no homologies with genes sequenced in other systems were detected.