The steady-state levels of microRNAs (miRNAs) and their activities are regulated by the post-transcriptional processes. It is known that 3′ ends of several miRNAs undergo post-dicing adenylation or uridylation. We isolated the liver-specific miR-122 from human hepatocytes and mouse livers. Direct analysis by mass spectrometry revealed that one variant of miR-122 has a 3′-terminal adenosine that is introduced after processing by Dicer. We identified GLD-2, which is a regulatory cytoplasmic poly(A) polymerase, as responsible for the 3′-terminal adenylation of miR-122 after unwinding of the miR-122/miR-122* duplex. In livers from GLD-2-null mice, the steady-state level of the mature form of miR-122 was specifically lower than in heterozygous mice, whereas no reduction of pre-miR-122 was observed, demonstrating that 3′-terminal adenylation by GLD-2 is required for the selective stabilization of miR-122 in the liver.

Macrocyclic peptides are an emerging class of therapeutics that can modulate protein–protein interactions. In contrast to the heavily automated high-throughput screening systems traditionally used for the identification of chemically synthesized small-molecule drugs, peptide-based macrocycles can be synthesized by ribosomal translation and identified using in vitro selection techniques, allowing for extremely rapid (hours to days) screening of compound libraries comprising more than 10¹³ different species. Furthermore, chemical modification of translated peptides and engineering of the genetic code have greatly expanded the structural diversity of the available peptide libraries. In this review, we discuss the use of these technologies for the identification of bioactive macrocyclic peptides, emphasizing recent developments.

Mammalian mitochondrial (mt) mRNAs have short poly(A) tails at their 3′ termini that are post-transcriptionally synthesized by mt poly(A) polymerase (PAP). The polyadenylation of mt mRNAs is known to be a key process needed to create UAA stop codons that are not encoded in mtDNA. In some cases, polyadenylation is required for the tRNA maturation by editing of its 3′ terminus. However, little is known about the functional roles the poly(A) tail of mt mRNAs plays in mt translation and RNA turnover. Here we show human mt PAP (hmtPAP) and human polynucleotide phosphorylase (hPNPase) control poly(A) synthesis in human mitochondria. Partial inactivation of hmtPAP by RNA interference using small interfering RNA in HeLa cells resulted in shortened poly(A) tails and decreased steady state levels of some mt mRNAs as well as their translational products. Moreover, knocking down hmtPAP generated markedly defective mt membrane potentials and reduced oxygen consumption. In contrast, knocking down hPNPase showed significantly extended poly(A) tails of mt mRNAs. These results demonstrate that the poly(A) length of human mt mRNAs is controlled by polyadenylation by hmtPAP and deadenylation by hPNPase, and polyadenylation is required for the stability of mt mRNAs.