Polyadenylation of ribosomal RNA in human cells

The addition of poly(A)-tails to RNA is a process common to almost all organisms. In eukaryotes, stable poly(A)-tails, important for mRNA stability and translation initiation, are added to the 3′ ends of most nuclear-encoded mRNAs, but not to rRNAs. Contrarily, in prokaryotes and organelles, polyadenylation stimulates RNA degradation. Recently, polyadenylation of nuclear-encoded transcripts in yeast was reported to promote RNA degradation, demonstrating that polyadenylation can play a double-edged role for RNA of nuclear origin. Here we asked whether in human cells ribosomal RNA can undergo polyadenylation. Using both molecular and bioinformatic approaches, we detected non-abundant polyadenylated transcripts of the 18S and 28S rRNAs. Interestingly, many of the post-transcriptionally added tails were composed of heteropolymeric poly(A)-rich sequences containing the other nucleotides in addition to adenosine. These polyadenylated RNA fragments are most likely degradation intermediates, as primer extension (PE) analysis revealed the presence of distal fragmented molecules, some of which matched the polyadenylation sites of the proximal cleavage products revealed by oligo(dT) and circled RT–PCR. These results suggest the presence of a mechanism to degrade ribosomal RNAs in human cells, that possibly initiates with endonucleolytic cleavages and involves the addition of poly(A) or poly(A)-rich tails to truncated transcripts, similar to that which operates in prokaryotes and organelles

LHC-like Proteins: The Guardians of Photosynthesis

The emergence of chlorophyll-containing light-harvesting complexes (LHCs) was a crucial milestone in the evolution of photosynthetic eukaryotic organisms. Light-harvesting chlorophyll-binding proteins form complexes in proximity to the reaction centres of photosystems I and II and serve as an antenna, funnelling the harvested light energy towards the reaction centres, facilitating photochemical quenching, thereby optimizing photosynthesis. It is now generally accepted that the LHC proteins evolved from LHC-like proteins, a diverse family of proteins containing up to four transmembrane helices. Interestingly, LHC-like proteins do not participate in light harvesting to elevate photosynthesis activity under low light. Instead, they protect the photosystems by dissipating excess energy and taking part in non-photochemical quenching processes. Although there is evidence that LHC-like proteins are crucial factors of photoprotection, the roles of only a few of them, mainly the stress-related psbS and lhcSR, are well described. Here, we summarize the knowledge gained regarding the evolution and function of the various LHC-like proteins, with emphasis on those strongly related to photoprotection. We further suggest LHC-like proteins as candidates for improving photosynthesis in significant food crops and discuss future directions in their research

Stable PNPase RNAi silencing: Its effect on the processing and adenylation of human mitochondrial RNA

Polynucleotide phosphorylase (PNPase) is a diverse enzyme, involved in RNA polyadenylation, degradation, and processing in prokaryotes and organelles. However, in human mitochondria, PNPase is located in the intermembrane space (IMS), where no mitochondrial RNA (mtRNA) is known to be present. In order to determine the nature and degree of its involvement in mtRNA metabolism, we stably silenced PNPase by establishing HeLa cell lines expressing PNPase short-hairpin RNA (shRNA). Processing and polyadenylation of mt-mRNAs were significantly affected, but, to different degrees in different genes. For instance, the stable poly(A) tails at the 3′ ends of COX1 transcripts were abolished, while COX3 poly(A) tails remained unaffected and ND5 and ND3 poly(A) extensions increased in length. Despite the lack of polyadenylation at the 3′ end, COX1 mRNA and protein accumulated to normal levels, as was the case for all 13 mt-encoded proteins. Interestingly, ATP depletion also altered poly(A) tail length, demonstrating that adenylation of mtRNA can be manipulated by indirect, environmental means and not solely by direct enzymatic activity. When both PNPase and the mitochondrial poly(A)-polymerase (mtPAP) were concurrently silenced, the mature 3′ end of ND3 mRNA lacked poly(A) tails but retained oligo(A) extensions. Furthermore, in mtPAP-silenced cells, truncated adenylated COX1 molecules, considered to be degradation intermediates, were present but harbored significantly shorter tails. Together, these results suggest that an additional mitochondrial polymerase, yet to be identified, is responsible for the oligoadenylation of mtRNA and that PNPase, although located in the IMS, is involved, most likely by indirect means, in the processing and polyadenylation of mtRNA