The topipotency of the germline is the full manifestation of the pluri- and multipotency of embryonic and adult stem cells, thus the germline and stem cells must share common mechanisms that guarantee their multipotentials in development. One of the few such known shared mechanisms is represented by Piwi proteins, which constitute one of the two subfamilies of the Argonaute protein family. Piwi proteins bind to Piwi-interacting RNAs (piRNAs) that are generally 26–31 nucleotides in length. Both Piwi proteins and piRNAs are most abundantly expressed in the germline. Moreover, Piwi proteins are expressed broadly in certain types of somatic stem/progenitor cells and other somatic cells across animal phylogeny. Recent studies indicate that the Piwi-piRNA pathway mediates epigenetic programming and post-transcriptional regulation, which may be responsible for its function in germline specification, gametogenesis, stem cell maintenance, transposon silencing, and genome integrity in diverse organisms.

Lin28 acts as a repressor of microRNA processing and as a post-transcriptional regulatory factor for a subset of mRNAs. Here we report that in human embryonic stem cells Lin28 facilitates the expression of the pivotal pluripotency factor Oct4 at the post-transcriptional level. We provide evidence that Lin28 binds Oct4 mRNA directly through high affinity sites within its coding region and that an interaction between Lin28 and RNA helicase A (RHA) may play a part in the observed regulation. We further demonstrate that decreasing RHA levels impairs Lin28-dependent stimulation of translation in a reporter system. Taken together with previous studies showing that RHA is required for efficient translation of a specific class of mRNAs, these findings suggest a novel mechanism by which Lin28 may affect target mRNA expression and represent the first evidence of post-transcriptional regulation of Oct4 expression by Lin28 in human embryonic stem cells.

Increasing evidence suggests that oxidative damage is associated with normal aging and several neurodegenerative diseases. Mild cognitive impairment (MCI), the phase between normal aging and early dementia, is a common problem in the elderly with many subjects going on to develop Alzheimer's disease (AD). Although increased DNA oxidation is observed in the AD brain, it is unclear when the oxidative damage begins. To determine if DNA oxidation occurs in the brain of subjects with MCI, we quantified multiple oxidized bases in nuclear and mitochondrial DNA isolated from frontal, parietal and temporal lobes and cerebellum of short post-mortem interval autopsies of eight amnestic patients with MCI and six age-matched control subjects using gas chromatography/ mass spectrometry with selective ion monitoring. We found statistically significant elevations (p < 0.05) of 8-hydroxyguanine, a widely studied biomarker of DNA damage, in MCI nuclear DNA from frontal and temporal lobe and in mitochondrial DNA from the temporal lobe compared with age-matched control subjects. Levels of 8-hydroxyadenine and 4,6-diamino-5-formamidopyrimidine were significantly elevated in nuclear DNA from all three neocortical regions in MCI. Statistically significant elevations of 4,6-diamino-5-formamidopyrimidine were also observed in mitochondrial DNA of MCI temporal, frontal and parietal lobes. These results suggest that oxidative damage to nuclear and mitochondrial DNA occurs in the earliest detectable phase of AD and may play a meaningful role in the pathogenesis of this disease.

SUMMARY Piwi proteins play essential roles in germline development, stem cell self-renewal, epigenetic regulation, and transposon silencing, and are implicated in oncogenesis [1-7]. Piwi proteins bind to a complex class of small non-coding RNAs called Piwi-Interacting RNAs (piRNAs) [8]. Mammalian Piwi proteins such as Mili are localized in the cytoplasm of spermatogenic cells, where they are associated with a germline-specific organelle called the nuage or its derivative, the chromatoid body, as well as with polysomes [9]. To investigate the molecular mechanisms mediated by Mili, we searched for Mili interacting proteins using co-immunoprecipitation and mass spectrometry. Here we report that Mili specifically interacts with Tudor Domain Containing Protein 1 (Tdrd1; a.k.a. Mouse Tudor Repeat 1, Mtr-1), a germline protein that contains multiple Tudor domains [10, 11]. This RNA-independent interaction is mediated through the N-terminal domain of Mili and the N-terminal region of Tdrd1 containing the Myeloid Nervy DEAF-1 (MYND) domain and first two Tudor domains. In addition, Mili positively regulates Tdrd1 expression at the mRNA level. Furthermore, Mili and Tdrd1 mutants share similar spermatogenic defects. However, Tdrd1, unlike Mili, is not required for piRNA biogenesis. Our results suggest that Mili interacts with Tdrd1 in the nuage and chromatoid body. This interaction does not contribute to piRNA biogenesis, but represents a regulatory mechanism critical for spermatogenesis.

Canalization, also known as developmental robustness, describes an organism's ability to produce the same phenotype despite genotypic variations and environmental influences 1,2. In Drosophila, Hsp90, the Trithorax group proteins, and transposon silencing have been implicated in canalization 3,4. Despite this, molecular mechanism underlying canalization remains elusive. Here, using an Drosophila eye-outgrowth assay sensitized by the dominant Kr Irregular facets-1 (Kr If-1), allele 3 , we show that the piRNA pathway, but not siRNA or miRNA pathways, is involved in canalization. Furthermore, we isolated a protein complex composed of Hsp90, Piwi, and the Hsp70/Hsp90 Organizing Protein Homolog (Hop), and demonstrated the function of this complex in canalization. Our data indicate that Hsp90 and Hop regulate the piRNA pathway via Piwi to mediate canalization. Moreover, they point to epigenetic silencing of the expression of existing genetic variants and the suppression of transposon-induced new genetic variation as two major mechanisms underlying piRNA pathway-mediated canalization. In both plants and animals, Hsp90 buffers against morphological changes induced either by genetic or non-genetic mechanisms, thereby promoting the robustness of the developmental programs that have been subjected to natural selection 5-8. However, under certain conditions, such as environmental stress, Hsp90 becomes overwhelmed, loosens its grip on canalization, and fails to repress the expression of genotype variants that have accumulated during evolution. The expressed phenotypes quickly become independent of Hsp90 deficiency, can be inherited in later generations, and could be subject to natural selection 3,5. In addition to Hsp90, maternally inherited epigenetic machineries also prevent expression of Users may view, print, copy, download and text and data-mine the content in such documents, for the purposes of academic research, subject always to the full Conditions of use:

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Degradation of extracellular matrix (ECM) underlies loss of cartilage tissue in osteoarthritis, a common disease for which no effective disease-modifying therapy currently exists. Here we describe BNTA, a small molecule with ECM modulatory properties. BNTA promotes generation of ECM components in cultured chondrocytes isolated from individuals with osteoarthritis. In human osteoarthritic cartilage explants, BNTA treatment stimulates expression of ECM components while suppressing inflammatory mediators. Intra-articular injection of BNTA delays the disease progression in a trauma-induced rat model of osteoarthritis. Furthermore, we identify superoxide dismutase 3 (SOD3) as a mediator of BNTA activity. BNTA induces SOD3 expression and superoxide anion elimination in osteoarthritic chondrocyte culture, and ectopic SOD3 expression recapitulates the effect of BNTA on ECM biosynthesis. These observations identify SOD3 as a relevant drug target, and BNTA as a potential therapeutic agent in osteoarthritis.

SUMMARYThe nuage is a germline-specific perinuclear structure that remains functionally elusive. Recently, the nuage in Drosophila was shown to contain two of the three PIWI proteins -Aubergine and Argonaute 3 (AGO3) -that are essential for germline development. The PIWI proteins bind to PIWI-interacting RNAs (piRNAs) and function in epigenetic regulation and transposon control. Here, we report a novel nuage component, PAPI (Partner of PIWIs), that contains a TUDOR domain and interacts with all three PIWI proteins via symmetrically dimethylated arginine residues in their N-terminal domain. In adult ovaries, PAPI is mainly cytoplasmic and enriched in the nuage, where it partially colocalizes with AGO3. The localization of PAPI to the nuage does not require the arginine methyltransferase dPRMT5 or AGO3. However, AGO3 is largely delocalized from the nuage and becomes destabilized in the absence of PAPI or dPRMT5, indicating that PAPI recruits PIWI proteins to the nuage to assemble piRNA pathway components. As expected, papi deficiency leads to transposon activation, phenocopying piRNA mutants. This further suggests that PAPI is involved in the piRNA pathway for transposon silencing. Moreover, AGO3 and PAPI associate with the P body component TRAL/ME31B complex in the nuage and transposon activation is observed in tral mutant ovaries. This suggests a physical and functional interaction in the nuage between the piRNA pathway components and the mRNA-degrading P-body components in transposon silencing. Overall, our study reveals a function of the nuage in safeguarding the germline genome against deleterious retrotransposition via the piRNA pathway.