Human neurodegenerative diseases possess the temporal hallmark of afflicting the elderly population. Hence, aging is among the most significant factors to impinge on disease onset and progression1, yet little is known of molecular pathways that connect these processes. Central to understanding this connection is to unmask the nature of pathways that functionally integrate aging, chronic maintenance of the brain and modulation of neurodegenerative disease. microRNAs (miRNA) are emerging as critical players in gene regulation during development, yet their role in adult-onset, age-associated processes are only beginning to be revealed. Here we report that the conserved miRNA miR-34 regulates age-associated events and long-term brain integrity in Drosophila, presenting such a molecular link between aging and neurodegeneration. Fly miR-34 expression is adult-onset, brain-enriched and age-modulated. Whereas miR-34 loss triggers a gene profile of accelerated brain aging, late-onset brain degeneration and a catastrophic decline in survival, miR-34 upregulation extends median lifespan and mitigates neurodegeneration induced by human pathogenic polyglutamine (polyQ) disease protein. Some of the age-associated effects of miR-34 require adult-onset translational repression of Eip74EF, an essential ETS domain transcription factor involved in steroid hormone pathways. These studies indicate that miRNA-dependent pathways may impact adult-onset, age-associated events by silencing developmental genes that later have a deleterious influence on adult life cycle and disease, and highlight fly miR-34 as a key miRNA with a role in this process

MicroRNAs (miRNAs) are 20-to~24-nucleotide (nt) small RNAs that impact a variety of biological processes, from development to age-associated events. To study the role of miRNAs in aging, studies have profiled the levels of miRNAs with time. However, evidence suggests that miRNAs show heterogeneity in length and sequence in different biological contexts. Here, by examining the expression pattern of miRNAs by Northern blot analysis, we found that Drosophila miRNAs show distinct isoform pattern changes with age. Surprisingly, an increase of some miRNAs reflects increased 29-O-methylation of select isoforms. Small RNA deep sequencing revealed a global increase of miRNAs loaded into Ago2, but not into Ago1, with age. Our data suggest increased loading of miRNAs into Ago2, but not Ago1, with age, indicating a mechanism for differential loading of miRNAs with age between Ago1 and Ago2. Mutations in Hen1 and Ago2, which lack 29-O-methylation of miRNAs, result in accelerated neurodegeneration and shorter life span, suggesting a potential impact of the ageassociated increase of 29-O-methylation of small RNAs on age-associated processes. Our study highlights that miRNA 29-O-methylation at the 39 end is modulated by differential partitioning of miRNAs between Ago1 and Ago2 with age and that this process, along with other functions of Ago2, might impact age-associated events in Drosophila.

An expanded (G4C2)30+ repeat within C9orf72 is the most prominent mutation in familial FTD and ALS. Through an unbiased, large-scale screen in (G4C2)49-expressing Drosophila we identify the CDC73/PAF1 complex (PAF1C), a transcriptional regulator of RNAPII, as a suppressor of G4C2-associated toxicity. Depletion of PAF1C reduces RNA and GR-dipeptide production from (G4C2)30+ transgenes. Interestingly, dPAF1C components, dPaf1 and dLeo1 appear selective for transcription of long, toxic repeat expansions, but not shorter, non-toxic expansions. In yeast, scPAF1C components regulate expression of both sense and anti-sense repeats. PAF1C is upregulated upon expression of (G4C2)30+ in flies and mice. hPaf1 is also upregulated in C9+ -derived cells and its heterodimer partner, hLeo1, binds C9 + repeat chromatin. In C9+ FTD, hPAF1 and hLEO1 are upregulated and their expression positively correlates with expression of repeat-containing C9orf72 transcripts. These data indicate that PAF1C activity is an important factor for transcription of the long, toxic repeat in C9+ FTD.

Amyotrophic lateral sclerosis (ALS) is a devastating, rapidly progressive disease leading to paralysis and death. Recently, intermediate length polyglutamine (polyQ) repeats of 27–33 in ATAXIN-2 (ATXN2), encoding the ATXN2 protein, were found to increase risk for ALS. In ATXN2, polyQ expansions of ≥34, which are pure CAG repeat expansions, cause spinocerebellar ataxia type 2. However, similar length expansions that are interrupted with other codons, can present atypically with parkinsonism, suggesting that configuration of the repeat sequence plays an important role in disease manifestation in ATXN2 polyQ expansion diseases. Here we determined whether the expansions in ATXN2 associated with ALS were pure or interrupted CAG repeats, and defined single nucleotide polymorphisms (SNPs) rs695871 and rs695872 in exon 1 of the gene, to assess haplotype association. We found that the expanded repeat alleles of 40 ALS patients and 9 long-repeat length controls were all interrupted, bearing 1–3 CAA codons within the CAG repeat. 21/21 expanded ALS chromosomes with 3CAA interruptions arose from one haplotype (GT), while 18/19 expanded ALS chromosomes with <3CAA interruptions arose from a different haplotype (CC). Moreover, age of disease onset was significantly earlier in patients bearing 3 interruptions vs fewer, and was distinct between haplotypes. These results indicate that CAG repeat expansions in ATXN2 associated with ALS are uniformly interrupted repeats and that the nature of the repeat sequence and haplotype, as well as length of polyQ repeat, may play a role in the neurological effect conferred by expansions in ATXN2.

Cotton fibers are single-cell trichomes derived from the outer integument cells of ovules. Recent studies showed that the rapid elongation of fibers was coordinated with the expression of a subset of fiber-specific genes. Here we report the use of filter arrays of cDNAs to identify genes preferentially expressed in cotton fibers. An array of 1536 clones was hybridized with cDNA probes prepared from wild type (fiber-containing) and fl mutant (naked) ovules at 5 days post-anthesis (DPA), respectively. Subsequent RT-PCR analyses of 14 genes identified ten that were highly transcribed in cotton fibers. Among them, an RD22-like protein (GhRDL ), a putative acyltransferase (GhACY ), a Fiddlehead homolog (GhFDH ), a serine carboxypeptidase-like protein (GhSCP ), two tubulin components (GhTUA6 and GhTUB1 ) and the previously reported gene encoding fiber protein E6, showed a fiberenriched expression pattern. The other three genes, including an actin (GhACT ), a putative cellulose synthase catalytic subunit (GhCesA-5 ) and a putative 24-sterol-C -methyltransferase (GhSMT ), were actively transcribed in fibers during the elongation stage, but their transcripts were also clearly present in other tissues. The possible roles of these proteins in cotton fiber development and growth are discussed. #