Extension of the Drosophila lifespan by overexpression of a protein repair methyltransferase.

Atypical protein isoaspartyl residues arise spontaneously during the aging process from the deamidation of protein asparaginyl residues and the isomerization of protein aspartyl residues. These abnormal residues are modified in cells by a strongly conserved protein carboxyl methyltransferase (PCMT) as a first step in a repair pathway. Because a decline in cellular repair mechanisms is hypothesized to contribute to senescence, we determined whether increased PCMT activity was correlated with enhanced longevity. Two ubiquitous promoters were used with the binary GAL4-UAS system to drive PCMT overexpression in Drosophila melanogaster. Flies expressing PCMT activity under the regulation of either the hsp70 or actin5C promoter had enzyme activities that were 3- or 7-fold higher, respectively, than control flies at 29 degrees C. Correlated with the observed increases in PCMT activities, such flies lived on average 32-39% longer than control flies. Lifespan extension was not observed at 25 degrees C with either hsp70- or actin5C-driven expression, indicating a temperature-dependent effect on longevity. We conclude that protein repair is an important factor in the determination of lifespan under certain environmental conditions. PCMT activity may become limiting under mild stress conditions that accelerate rates of protein damage.

Translation of a unique transcript for protein isoaspartyl methyltransferase in haploid spermatids: implications for protein storage and repair.

The mammalian testis contains high levels of a protein, L-isoaspartyl (D-aspartyl) O-methyltransferase (PIMT), postulated to play a role in the repair of age-damaged proteins. To examine the regulation of PIMT concentrations during the development of spermatozoa, poly(A)+ RNA was isolated from purified populations of pachytene spermatocytes and round spermatids. Northern blot analysis revealed that a unique 1.1-1.3 kb PIMT transcript is present in preparations of round spermatid and pachytene spermatocyte poly (A)+ RNA. The concentration of this small PIMT transcript is at least four times higher in mRNA isolated from round spermatids than in mRNA isolated from pachytene spermatocytes, indicating that the PIMT gene is actively transcribed during the haploid phase of spermatogenesis. The germ cell-specific PIMT transcripts are distributed between the polysomal fraction and the nonpolysomal fractions of testis RNA, suggesting that translational controls also contribute to the high concentrations of PIMT in mammalian sperm. PIMT function is not essential for spermatogenesis because the testes from transgenic mice lacking PIMT activity have normal levels of protamine transcripts, and because functional sperm can be recovered from the cauda epididymis. The protein repair function of the PIMT may be more important in maintaining the fertilization competence of translationally-inactive mature sperm during the prolonged period of epididymal transit and storage in the male reproductive tract.