Characteristics of the endocrine system in Drosophila melanogaster 1 (2)gl mutants, differing in time of death]

A comparative study of the state of the endocrine system and one of the larval organs, the salivary gland, has been carried out in larvae homozygous for the 1(2)gl gene. They differ in time of death (death at the third larval instar--larval allele, and at the prepupal state--prepupal allele). It is shown that homozygotes for the larval and prepupal allele have underdeveloped prothoracal glands. Corpora allata in homozygotes for the larval allele does not differ from the norm. Corpora allata in homozygotes for the prepupal allele is decreased proportionally to the decrease of prothoracal glands. A decrease of gland size is due to a decrease of the volume of cell but not to their number; this decrease is accompanied by the decrease of their relative DNA content. Salivary glands in homozygotes are reduced and comprise 80% of the normal size in homozygotes for the prepupal allele and 50% for homozygotes for the larval allele. Polyteny level in the salivary gland nuclei is much decreased as compared with the normal level. DNA level is more reduced in larvae homozygous for the larval allele.     

Why,when and how does the poly(A) tail shorten during mRNA translation?

• 1.1. The length of the poly(A) tail at the 3'-end of mRNA may control protein synthesis by bringing the 3'-end in close proximity to the 5'-end of the noncoding region as well as increasing the duration of mRNA translation by its binding to the poly(A) binding protein.
• 2.2. The rate-limiting step in the decay of the body of the message is the shortening of a long poly(A) tail during mRNA translation. The shortening of the poly(A) tail occurs during pre-elongation in the protein synthesis cycle.
• 3.3. The shortening of the poly(A) tail during mRNA translation may not involve RNase activity, however poly(A) binding protein seems to play a role, at least in part, in shortening of the poly(A) tail.

     

Determinants of mRNA stability in Dictyostelium discoideum amoebae: differences in poly(A) tail length, ribosome loading, and mRNA size cannot account for the heterogeneity of mRNA decay rates.

As an approach to understanding the structures and mechanisms which determine mRNA decay rates, we have cloned and begun to characterize cDNAs which encode mRNAs representative of the stability extremes in the poly(A)+ RNA population of Dictyostelium discoideum amoebae. The cDNA clones were identified in a screening procedure which was based on the occurrence of poly(A) shortening during mRNA aging. mRNA half-lives were determined by hybridization of poly(A)+ RNA, isolated from cells labeled in a 32PO4 pulse-chase, to dots of excess cloned DNA. Individual mRNAs decayed with unique first-order decay rates ranging from 0.9 to 9.6 h, indicating that the complex decay kinetics of total poly(A)+ RNA in D. discoideum amoebae reflect the sum of the decay rates of individual mRNAs. Using specific probes derived from these cDNA clones, we have compared the sizes, extents of ribosome loading, and poly(A) tail lengths of stable, moderately stable, and unstable mRNAs. We found (i) no correlation between mRNA size and decay rate; (ii) no significant difference in the number of ribosomes per unit length of stable versus unstable mRNAs, and (iii) a general inverse relationship between mRNA decay rates and poly(A) tail lengths. Collectively, these observations indicate that mRNA decay in D. discoideum amoebae cannot be explained in terms of random nucleolytic events. The possibility that specific 3'-structural determinants can confer mRNA instability is suggested by a comparison of the labeling and turnover kinetics of different actin mRNAs. A correlation was observed between the steady-state percentage of a given mRNA found in polysomes and its degree of instability; i.e., unstable mRNAs were more efficiently recruited into polysomes than stable mRNAs. Since stable mRNAs are, on average, "older" than unstable mRNAs, this correlation may reflect a translational role for mRNA modifications that change in a time-dependent manner. Our previous studies have demonstrated both a time-dependent shortening and a possible translational role for the 3' poly(A) tracts of mRNA. We suggest, therefore, that the observed differences in the translational efficiency of stable and unstable mRNAs may, in part, be attributable to differences in steady-state poly(A) tail lengths.