Summary DNA mismatch repair (MMR) ensures replication fidelity by correcting mismatches generated during DNA replication. Although human MMR has been reconstituted in vitro, how MMR occurs in vivo is unknown. Here, we show that an epigenetic histone mark, H3K36me3, is required in vivo to recruit the mismatch recognition protein hMutSα (hMSH2-hMSH6) onto chromatin through direct interactions with the hMSH6 PWWP domain. The abundance of H3K36me3 in G1 and early S phases ensures that hMutSα is enriched on chromatin before mispairs are introduced during DNA replication. Cells lacking the H3K36 tri-methyltransferase SETD2 display microsatellite instability (MSI) and an elevated spontaneous mutation frequency, characteristic of MMR-deficient cells. This work reveals that a histone mark regulates MMR in human cells and explains the long-standing puzzle of MSI-positive cancer cells that lack detectable mutations in known MMR genes.

The miR-15/107 group of microRNA (miRNA) genes is increasingly appreciated to serve key functions in humans. These miRNAs regulate gene expression involved in cell division, metabolism, stress response, and angiogenesis in vertebrate species. The miR-15/107 group has also been implicated in human cancers, cardiovascular disease, and neurodegenerative diseases including Alzheimer's disease. Here, we provide an overview of (1) the evolution of miR-15/107 group member genes, (2) the expression levels of the miRNAs in mammalian tissues, (3) evidence for overlapping gene regulatory functions by the different miRNAs, (4) the normal biochemical pathways regulated by miR-15/107 group miRNAs, and (5) the roles played by these miRNAs in human diseases. Membership in this group is defined on the basis of sequence similarity near the mature miRNAs' 5′ end: all include the sequence AGCAGC. Phylogeny of this group of miRNAs is incomplete so a definitive taxonomic classification (for example, designation as a "superfamily") is currently not possible. While all vertebrates studied to date express miR-15a, -15b, -16, -103, and -107, mammals alone are known to express miR-195, -424, -497, -503, and -646. Multiple different miRNAs in the miR-15/107 group are expressed at moderate-to-high levels in human tissues. We present data on the expression of all known miR-15/107 group members in human cerebral cortical gray and white matter using new miRNA profiling microarrays. There is extensive overlap in the mRNAs targeted by miR-15/107 group members. We show new data from cultured H4 cancer cells that demonstrate similarities in mRNAs targeted by miR-16 and miR-103, and also support the importance of the mature miRNAs' 5′ seed region in mRNA target recognition. In conclusion, the miR-15/107 group of miRNA genes is a fascinating topic of study for evolutionary biologists, miRNA biochemists, and clinically-oriented translational researchers alike.

Patients with Alzheimer's disease (AD) exhibit higher levels of 8-oxo-guanine (8-oxoG) DNA lesions in their brain, suggesting a reduced or defective 8-oxoG repair. To test this hypothesis, this study investigated 14 AD patients and 10 age-matched controls for mutations of the major 8-oxoG removal gene OGG1. Whereas no alterations were detected in any control samples, four AD patients exhibited mutations in OGG1, two carried a common single base (C796) deletion that alters the carboxyl terminal sequence of OGG1, and the other two had nucleotide alterations leading to single amino acid substitutions. In vitro biochemical assays revealed that the protein encoded by the C796-deleted OGG1 completely lost its 8-oxoG glycosylase activity, and that the two single residue-substituted OGG1 proteins showed a significant reduction in the glycosylase activity. These results were consistent with the fact that nuclear extracts derived from a limited number of AD patients with OGG1 mutations exhibited greatly reduced 8-oxoG glycosylase activity compared with age-matched controls and AD patients without OGG1 alterations. Our findings suggest that defects in OGG1 may be important in the pathogenesis of AD in a significant fraction of AD patients and provide new insight into the molecular basis for the disease.

Somatic mutations on glycine 34 of histone H3 (H3G34) cause pediatric cancers, but the underlying oncogenic mechanism remains unknown. We demonstrate that substituting H3G34 with arginine, valine, or aspartate (H3G34R/V/D), which converts the non-side chain glycine to a large side chain-containing residue, blocks H3 lysine 36 (H3K36) dimethylation and trimethylation by histone methyltransferases, including SETD2, an H3K36-specific trimethyltransferase. Our structural analysis reveals that the H3 "G33-G34" motif is recognized by a narrow substrate channel, and that H3G34/R/V/D mutations impair the catalytic activity of SETD2 due to steric clashes that impede optimal SETD2-H3K36 interaction. H3G34R/V/D mutations also block H3K36me3 from interacting with mismatch repair (MMR) protein MutSα, preventing the recruitment of the MMR machinery to chromatin. Cells harboring H3G34R/V/D mutations display a mutator phenotype similar to that observed in MMR-defective cells. Therefore, H3G34R/V/D mutations promote genome instability and tumorigenesis by inhibiting MMR activity.

Eight-hydroxy-2'-deoxyguanosine (8-OHdG) is increased in the brain in late-stage Alzheimer's disease (LAD) and mild cognitive impairment (MCI). To determine if decreased base-excision repair contributes to these elevations, we measured oxoguanine glycosylase 1 (OGG1) protein and incision activities in nuclear and mitochondrial fractions from frontal (FL), temporal (TL), and parietal (PL) lobes from 8 MCI and 7 LAD patients, and 6 age-matched normal control (NC) subjects. OGG1 activity was significantly (P<0.05) decreased in nuclear specimens of FL, TL, and PL in MCI and LAD and in mitochondria from LAD FL and TL and MCI TL. Nuclear OGG1 protein was significantly decreased in LAD FL and MCI and LAD PL. No differences in mitochondrial OGG1 protein levels were found. Overall, our results suggest that decreased OGG1 activity occurs early in the progression of AD, possibly mediated by 4-hydroxynonenal inactivation and may contribute to elevated 8-OHdG in the brain in MCI and LAD.

Background: Expansion of CAG/CTG repeats causes familial neurological disorders, but the molecular basis is unknown. Results: DNA polymerases ␤ and ␦ effectively incorporate nucleotides to a CAG/CTG hairpin primer, leading to hairpin retention. Conclusion: Coordinated actions by polymerases ␤ and ␦ on hairpin primers during DNA synthesis promotes CAG/CTG repeat expansions. Significance: The work discovers a novel mechanism for CAG/CTG repeat expansions.

MicroRNAs (miRNAs) target mRNAs in human cells via complex mechanisms that are still incompletely understood. Using anti-Argonaute (anti-AGO) antibody co-immunoprecipitation, followed by microarray analyses and downstream bioinformatics, ‘RIP-Chip’ experiments enable direct analyses of miRNA targets. RIP-Chip studies (and parallel assessments of total input mRNA) were performed in cultured H4 cells after transfection with miRNAs corresponding to the miR-15/107 gene group (miR-103, miR-107, miR-16 and miR-195), and five control miRNAs. Three biological replicates were run for each condition with a total of 54 separate human Affymetrix Human Gene 1.0 ST array replicates. Computational analyses queried for determinants of miRNA:mRNA binding. The analyses support four major findings: (i) RIP-Chip studies correlated with total input mRNA profiling provides more comprehensive information than using either RIP-Chip or total mRNA profiling alone after miRNA transfections; (ii) new data confirm that miR-107 paralogs target coding sequence (CDS) of mRNA; (iii) biochemical and computational studies indicate that the 3′ portion of miRNAs plays a role in guiding miR-103/7 to the CDS of targets; and (iv) there are major sequence-specific targeting differences between miRNAs in terms of CDS versus 3′-untranslated region targeting, and stable AGO association versus mRNA knockdown. Future studies should take this important miRNA-to-miRNA variability into account.

MicroRNAs (miRNAs) are critical post-transcriptional regulators and are derived from hairpin-shaped primary transcripts via a series of processing steps. However, how the production of individual miRNAs is regulated remains largely unknown. Similarly, loss or overexpression of the key mismatch repair protein MutLα (MLH1-PMS2 heterodimer) leads to genome instability and tumorigenesis, but the mechanisms controlling MutLα expression are unknown. Here we demonstrate in vitro and in vivo that MLH1 and miR-422a participate in a feedback loop that regulates the level of both molecules. Using a defined in-vitro miRNA processing system, we show that MutLα stimulates the conversion of pri-miR-422a to pre-miR-422a, as well as the processing of other miRNAs tested, implicating MutLα as a general stimulating factor for miRNA biogenesis. This newly identified MutLα function requires its ATPase and pri-miRNA binding activities. In contrast, miR-422a downregulates MutLα levels by suppressing MLH1 expression through base pairing with the MLH1 3′-untranslated region. A model depicting this feedback mechanism is discussed.