Multifaceted Microcephaly-Related Gene MCPH1

MCPH1, or BRIT1, is often mutated in human primary microcephaly type 1, a neurodevelopmental disorder characterized by a smaller brain size at birth, due to its dysfunction in regulating the proliferation and self-renewal of neuroprogenitor cells. In the last 20 years or so, genetic and cellular studies have identified MCPH1 as a multifaceted protein in various cellular functions, including DNA damage signaling and repair, the regulation of chromosome condensation, cell-cycle progression, centrosome activity and the metabolism. Yet, genetic and animal model studies have revealed an unpredicted essential function of MPCH1 in gonad development and tumorigenesis, although the underlying mechanism remains elusive. These studies have begun to shed light on the role of MPCH1 in controlling various pathobiological processes of the disorder. Here, we summarize the biological functions of MCPH1, and lessons learnt from cellular and mouse models of MCPH1.

TRIM29 in Cutaneous Squamous Cell Carcinoma

Tripartite motif (TRIM) proteins play important roles in a wide range of cell physiological processes, such as signal transduction, transcriptional regulation, innate immunity, and programmed cell death. TRIM29 protein, encoded by the ATDC gene, belongs to the RING-less group of TRIM protein family members. It consists of four zinc finger motifs in a B-box domain and a coiled-coil domain, and makes use of the B-box domain as E3 ubiquitin ligase in place of the RING. TRIM29 was found to be involved in the formation of homodimers and heterodimers in relation to DNA binding; additional studies have also demonstrated its role in carcinogenesis, DNA damage signaling, and the suppression of radiosensitivity. Recently, we reported that TRIM29 interacts with keratins and FAM83H to regulate keratin distribution. Further, in cutaneous SCC, the expression of TRIM29 is silenced by DNA methylation, leading to the loss of TRIM29 and promotion of keratinocyte migration. This paper reviews the role of TRIM family proteins in malignant tumors, especially the role of TRIM29 in cutaneous SCC.

Diabetic Pneumopathy–A New Diabetes-Associated Complication: Mechanisms, Consequences and Treatment Considerations

Patients with diabetes are over-represented among the total cases reported with “idiopathic” pulmonary fibrosis (IPF). This raises the question, whether this is an association only or whether diabetes itself can cause pulmonary fibrosis. Recent studies in mouse models of type 1 and type 2 diabetes demonstrated that diabetes causes pulmonary fibrosis. Both types of diabetes trigger a cascade, starting with increased DNA damage, an impaired DNA repair, and leading to persistent DNA damage signaling. This response, in turn, induces senescence, a senescence-associated-secretory phenotype (SASP), marked by the release of pro-inflammatory cytokines and growth factors, finally resulting in fibrosis. Restoring DNA repair drives fibrosis into remission, thus proving causality. These data can be translated clinically to patients with type 2 diabetes, characterized by long-term diabetes and albuminuria. Hence there are several arguments, to substitute the term “idiopathic” pulmonary fibrosis (IPF) in patients with diabetes (and exclusion of other causes of lung diseases) by the term “diabetes-induced pulmonary fibrosis” (DiPF). However, future studies are required to establish this term and to study whether patients with diabetes respond to the established therapies similar to non-diabetic patients.

Collateral Victim or Rescue Worker?—The Role of Histone Methyltransferases in DNA Damage Repair and Their Targeting for Therapeutic Opportunities in Cancer

Disrupted DNA damage signaling greatly threatens cell integrity and plays significant roles in cancer. With recent advances in understanding the human genome and gene regulation in the context of DNA damage, chromatin biology, specifically biology of histone post-translational modifications (PTMs), has emerged as a popular field of study with great promise for cancer therapeutics. Here, we discuss how key histone methylation pathways contribute to DNA damage repair and impact tumorigenesis within this context, as well as the potential for their targeting as part of therapeutic strategies in cancer.

Ataxia with Ocular Apraxia Type 1 (AOA1) (APTX, W279* Mutation): Neurological, Neuropsychological, and Molecular Outlining of a Heterogenous Phenotype in Four Colombian Siblings

Hereditary ataxias are a group of devastating neurological disorders that affect coordination of gait and are often associated with poor coordination of hands, speech, and eye movements. Ataxia with Ocular Apraxia type 1 (AOA1) (OMIM: 606350.0006) is characterized by slowly progressive symptoms of childhood-onset and pathogenic mutations in APTX; the only known cause underpinning AOA1. APTX encodes the protein Aprataxin, composed of three domains sharing homology with proteins involved in DNA damage, signaling, and repair. We present four siblings from an endogamic family in a rural, isolated town of Colombia with ataxia and ocular apraxia of childhood-onset and confirmed molecular diagnosis of AOA1, homozygous for the W279* p.Trp279Ter mutation. We predicted the mutated APTX with Alpha Fold to demonstrate the effects of this stop-gain mutation that deletes three beta helices encoded by amino acid 270 to 339 rescinding the C2H2-type zinc fingers (Znf) (C2H2 Znf) DNA-binding and DNA-repair domain and the whole tridimensional structure of the APTX. All siblings exhibited different ages of onset (4, 6, 8, and 11 y/o) and heterogeneous patterns of dysarthria (ranging from absence to mild-moderate dysarthria). Neuropsychological evaluation showed no neurocognitive impairment in three siblings, but one sibling showed temporospatial disorientation, semantic and phonologic fluency impairment, episodic memory affection, constructional apraxia, moderate anomia, low executive function, and symptoms of depression. This heterogeneous phenotype suggests genetic interactions can shape the natural history of AOA1. To our knowledge, this report represents the most extensive series of siblings affected with AOA1 in Latin America, and the genetic analysis completed adds important knowledge to outline this family’s disease and general complex phenotype of hereditary ataxias.

CBIO-08. ENDOGENOUS DNA DOUBLE STRAND BREAKS ACTIVATE HETEROGENOUS DNA DAMAGE SIGNALING IN IDH1/2 MUTANT GLIOMAS

Isocitrate dehydrogenase 1/2 (IDH1/2) mutations are common in astrocytic glioma and are frequently coupled with TP53 and ATRX mutations. Collectively, these alterations cause genomic instability leading to high basal DNA double strand breaks (DSBs). Understanding how IDH/TP53/ATRX mutant cells process endogenous DSBs may help exploit inhibitors of DNA damage response (DDR) for the treatment of patients with IDH mutant gliomas. Through systematic effort to uncover the mechanisms involved in repair of endogenous DSBs in IDH1/2 mutant GBMs, we have discovered that high basal phosphorylated DNA-PK (p-DNA-PK) was characteristic of an IDH1/TP53/ATRX mutant GBM164 patient derived xenograft (PDX) but not in another IDH1 mutant GBM196 PDX. Immunofluorescence (IF) studies in patient specimen from which GBM164 was derived showed that p-DNA-PK co-localized with g-H2AX, 53BP1 or H4K20me2 (but not p-RPA) the known surrogates of DSBs. In contrast, p-DNA-PK was absent in the patient specimen from which GBM196 was derived, which otherwise had equally intense g-H2AX immunostaining colocalized with p-RPA. An independent IF study involving 11 IDH1 wild-type (WT) and 11 IDH1 mutant GBM patient samples, the p-DNA-PK was observed in 3 (27%) of 11 IDH1 mutant samples while IDH1 WT tumors were negative for p-DNA-PK. A telomere specific fluorescence in situ hybridization (Tel-FISH) confirmed elevated alternative lengthening of telomere (ALT) activity in GBM196 (but not in GBM164) indicative of HR proficiency. Consistently, HR related genes, including BRCA1 and MRE11A, were found upregulated in ALT-positive GBM196 as compared to those in GBM164. Interestingly, ALT+ GBM196 cells were highly vulnerable to inhibitors of ATM and ATR pathways. In conclusion, IDH1/TP53/ATRX mutant gliomas can be subdivided into HR-mediated ALT-positive group, which repairs the endogenous DSBs by HR (e.g. GBM196) and an ALT-negative/p-DNA-PK group, which repairs DSBs by c-NHEJ (e.g. GBM164) and this subdivision can be developed as a prescient biomarker of sensitivity to DDR inhibitors.

NAD-Linked Metabolism and Intervention in Short Telomere Syndromes and Murine Models of Telomere Dysfunction

Telomeres are specialized nucleoprotein structures that form protective caps at the ends of chromosomes. Short telomeres are a hallmark of aging and a principal defining feature of short telomere syndromes, including dyskeratosis congenita (DC). Emerging evidence suggests a crucial role for critically short telomere-induced DNA damage signaling and mitochondrial dysfunction in cellular dysfunction in DC. A prominent factor linking nuclear DNA damage and mitochondrial homeostasis is the nicotinamide adenine dinucleotide (NAD) metabolite. Recent studies have demonstrated that patients with DC and murine models with critically short telomeres exhibit lower NAD levels, and an imbalance in the NAD metabolome, including elevated CD38 NADase and reduced poly (ADP-ribose) polymerase and SIRT1 activities. CD38 inhibition and/or supplementation with NAD precursors reequilibrate imbalanced NAD metabolism and alleviate mitochondrial impairment, telomere DNA damage, telomere dysfunction-induced DNA damage signaling, and cellular growth retardation in primary fibroblasts derived from DC patients. Boosting NAD levels also ameliorate chemical-induced liver fibrosis in murine models of telomere dysfunction. These findings underscore the relevance of NAD dysregulation to telomeropathies and demonstrate how NAD interventions may prove to be effective in combating cellular and organismal defects that occur in short telomere syndromes.

Persistent DNA damage signaling and DNA polymerase theta promote broken chromosome segregation

Cycling cells must respond to DNA double-strand breaks (DSBs) to avoid genome instability. Missegregation of chromosomes with DSBs during mitosis results in micronuclei, aberrant structures linked to disease. How cells respond to DSBs during mitosis is incompletely understood. We previously showed that Drosophilamelanogaster papillar cells lack DSB checkpoints (as observed in many cancer cells). Here, we show that papillar cells still recruit early acting repair machinery (Mre11 and RPA3) and the Fanconi anemia (FA) protein Fancd2 to DSBs. These proteins persist as foci on DSBs as cells enter mitosis. Repair foci are resolved in a stepwise manner during mitosis. DSB repair kinetics depends on both monoubiquitination of Fancd2 and the alternative end-joining protein DNA polymerase θ. Disruption of either or both of these factors causes micronuclei after DNA damage, which disrupts intestinal organogenesis. This study reveals a mechanism for how cells with inactive DSB checkpoints can respond to DNA damage that persists into mitosis.

DNA damage signaling regulates cohesin stabilization and promotes meiotic chromosome axis morphogenesis

A hallmark of meiosis is the reorganization of chromosomes as linear arrays of chromatin loops around a chro- mosome axis comprised of cohesins and regulatory proteins. Defective axis morphogenesis impairs homolog pairing, synapsis, and recombination. We find that axis assembly in C. elegans is promoted by DNA Damage Response (DDR) signaling activated at meiotic entry. Central to this regulation is downregulation of the cohesin release factor WAPL-1 by the DDR transducer kinase ATM-1, which is activated by the meiotic kinase CHK- 2. Additional cohesin regulators, including ECO-1 and PDS-5, also contribute to stabilizing axis-associated cohesins. We find that downregulation of WAPL by ATM also promotes cohesin enrichment at DNA damage foci in cultured mammalian cells. Our findings reveal that the DDR and Wapl play conserved roles in cohesin regulation in meiotic prophase and proliferating cells.

Mre11-Rad50 oligomerization promotes DNA double-strand break repair

The conserved Mre11-Rad50 (MR) complex is crucial for the detection, signaling, end tethering and processing of DNA double-strand breaks (DSBs). While it was known for decades that MR foci formation at DSBs accompanies repair, the underlying molecular assembly mechanisms and functional implications remained unclear. Combining pathway reconstitution in electron microscopy, biochemical assays and genetic studies, we show that S. cerevisiae MR oligomerizes via a conserved Rad50 beta-sheet to higher-order assemblies, which bind DNA with positive cooperativity. We designed Rad50 point mutants with enhanced or disrupted MR oligomerization, and demonstrate that MR oligomerization facilitates foci formation, DNA damage signaling and repair in vivo. MR oligomerization does not affect its exonuclease activity but drives endonucleolytic cleavage at multiple sites on the 5'-terminated DNA strand near DSBs. Interestingly, mutations in the human Rad50 beta-sheet are linked to hereditary cancer predisposition and our findings might provide new insights into their potential role in chemoresistance.