Javier Pena Diaz

Javier Pena Diaz

Guest researcher

Primary fields of research

Mismatch Repair roles in disease;
the Good, the Mischievous and the Ugly.
Originally, the DNA repair system mismatch repair (MMR) was described in bacteria as a system that prevented accumulation of mutations. In this capacity, it was later found to maintain genomic stability in a range of organisms. The clinical relevance of these studies was revealed when in humans, deficiencies in MMR were found to cause hypermutating phenotypes and predisposition to development of several cancers, including cancers of the colon and endometrium. The mechanism of action and the factors involved were identified using biochemical approaches and MMR was placed as one of the main safeguards of genome stability. Surprisingly, it was later found that MMR is also associated with mutagenic DNA transactions at the immunoglobulin locus. Here, the high fidelity function of MMR is corrupted and introduces rather than prevents mutations. We coined the term non-canonical MMR (ncMMR) to describe this mutagenic pathway. At the immunoglobulin locus, this mischievous version of MMR contributes in a beneficial way to the creation of the antibody repertoire needed, but ncMMR contributes also to disease when functioning at different loci. This is the case of ncMMR activities at complex DNA repetitive sequences that can form secondary structures. In particular, ncMMR promotes instability at trinucleotide repeats that is the underlying cause of several neurological disorders.

My group is interesting in understanding how a high fidelity DNA repair mechanisms turns into a mutagenic one. We focus on identifying the factors that may affect this pathway choice and analysing the function of ncMMR at DNA sequences that be susceptible to its mutagenic action such as trinucleotide repeats and telomeres.

Current research

The MMR interactome
Several interactome studies have revealed a complex landscape of interactions between MMR proteins and proteins involved in replication, chromatin maintenance, factors which are found at ssDNA tracks, factors that contribute to repair of dsbreaks and some molecular tools such as the helicases. Up to date, the relevance of some of these interactions remains largely unknown. Our goal is to determine whether any these factors play a role promoting the mutagenic activity of ncMMR.

Chromatin dynamics and MMR. Scheme of the processes affecting chromatin status where MMR is involved. Coloured proteins are those found as MMR interactors in various studies. In red, proteins found in our studies, green; Cannavo E et al. [1], blue; Li F et al. [2], magenta; Liu Y et al. [3], yellow; Wang Y et al. [4] and purple; Saydam N et al. [5]. The X-ray structures depicted were obtained from RCSB PDB Protein Data Bank, the cryo-EM structure of the ORC-Cdc6-Cdt1-Mcm 2-7 complex was obtained from Sun J et al. [6].


MMR in trinucleotide repeat expansion associated with neurological disorders
An increasing number of neurological disorders are found to be caused by expansion of certain repetitive sequences. These repetitive sequences have a define length in healthy individuals and an expanded length above a threshold which has pathological consequences. The most common of the repeats associated to disease are trinucleotide repeats (TNR) and the clinical manifestations vary depending on the gene affected. On the other hand, the mechanisms that promote the instability of these sequences seem to be conserved. Instability is believed to be caused by the formation of mutagenic slipped-DNA structures at these sequences and aberrant processing by DNA repair machineries (such as MMR). Our current focus of research is CAG repeat expansion (associated with Huntington’s disease and several spinocerebellar ataxias), and the involvement of the DNA repair machinery, mismatch repair (MMR). CAG repeats in coding regions of the gene encode for glutamine tracts (poly-Q). Expanded CAG repeats lead to longer poly-Q tracts known to become unstable and promote formation of protein aggregates typical of disease (e.g. huntingtin protein aggregates in Huntington’s disease patients). Our goal is to identify the factors and mechanisms that mediate MMR-dependent TNR expansion.

Short list and location of expandable repeats responsible for human diseases. (A) The sequence and location within a generic gene of expandable repeats that cause human diseases are shown, and the associated diseases are listed. Disease-associated unstable repeats affected by mismatch repair are depicted in red. BPES, blepharophimosis, ptosis and epicanthus inversus syndrome; CCD, cleidocranial dysplasia; CCHS, congenital central hypoventilation syndrome; DM, myotonic dystrophy; DRPLA, dentatorubral–pallidoluysian atrophy; EPM1, progressive myoclonic epilepsy 1; FRAXA, fragile X syndrome; FRAXE, fragile X mental retardation associated with FRAXE site; FRDA, Friedreich's ataxia; FXTAS, fragile X tremor and ataxia syndrome; HD, Huntington's disease; HDL2, Huntington's-disease-like 2; HFG, hand–foot–genital syndrome; HPE5, holoprosencephaly 5; ISSX, X-linked infantile spasm syndrome; MRGH, mental retardation with isolated growth hormone deficiency; OPMD, oculopharyngeal muscular dystrophy; SBMA, spinal and bulbar muscular atrophy; SCA, spinocerebellar ataxia; SPD, synpolydactyly. Red circles highlight diseases known to be associated with ncMMR mutagenic activity. Adapted from Mirkin S.M [7]. (B) Comparison between post-morten brain form a healthy individual and a HD affected patient shows atrophy of the caudate and increase in size of lateral ventricles. The Immunofluorescence images show distribution of aggregates in the cellular body of striatal neurons stained against huntingtin (green) and tubulin (red).Adapted from Li H. et al. [8].


MMR at telomeres
One of the requirements to achieve cell immortality is to preserve or maintain the end of the chromosomes, the so-called telomeres. Most cancer cells achieve this goal by re-activation of the enzyme telomerase but in up to 15% of all cancers the telomeres are maintain in a telomerase independent manner by a mechanism relying on recombination between telomeres. This alternative lengthening of telomeres (ALT) pathway is prevalent among cancers with poor prognosis such as glioblastomas and sarcomas.  How cells engage and regulate the ALT phenotype remains largely unknown. One of the mechanisms proposed to affect this choice is mismatch repair (MMR). Our research focuses in analysing the influence of MMR in telomere maintenance and the interference of MMR with other DNA repair enzymes that modulate telomere homeostasis.

MMR proteins are enriched at telomeres in ALT cancer cells. ImmunoFISH analysis of localization patterns of MLH1 (blue), MSH2 (red) and telomeres (green) in U2OS (ALT+) and HeLa (telomerase+) cells showing enrichment of the mismatch repair proteins at telomeres in the ALT cancer cell.

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