XRCC2 R188H (rs3218536), XRCC3 T241M (rs861539) and R243H (rs77381814) single nucleotide polymorphisms in cervical cancer risk

Human Papillomavirus (HPV) is the main cause of cervical cancer and its precursor lesions. Transformation may be induced by several mechanisms, including oncogene activation and genome instability. Individual differences in DNA damage recognition and repair have been hypothesized to influence cervical cancer risk. The aim of this study was to evaluate whether the double strand break gene polymorphisms XRCC2 R188H G>A (rs3218536), XRCC3 T241M C>T (rs861539) and R243H G>A (rs77381814) are associated to cervical cancer in Argentine women. A case control study consisting of 322 samples (205 cases and 117 controls) was carried out. HPV DNA detection was performed by PCR and genotyping of positive samples by EIA (enzyme immunoassay). XRCC2 and 3 polymorphisms were determined by pyrosequencing. The HPV-adjusted odds ratio (OR) of XRCC2 188 GG/AG genotypes was OR = 2.4 (CI = 1.1-4.9, p = 0.02) for cervical cancer. In contrast, there was no increased risk for cervical cancer with XRCC3 241 TT/CC genotypes (OR = 0.48; CI = 0.2-1; p = 0.1) or XRCC3 241 CT/CC (OR = 0.87; CI = 0.52-1.4; p = 0.6). Regarding XRCC3 R243H, the G allele was almost fixed in the population studied. In conclusion, although the sample size was modest, the present data indicate a statistical association between cervical cancer and XRCC2 R188H polymorphism. Future studies are needed to confirm these findings.Fil: Perez, Luis Orlando. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico CONICET- La Plata. Instituto de Genética Veterinaria "Ing. Fernando Noel Dulout". Universidad Nacional de La Plata. Facultad de Ciencias Veterinarias. Instituto de Genética Veterinaria; ArgentinaFil: Crivaro, Andrea Natalia. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico CONICET- La Plata. Instituto de Genética Veterinaria "Ing. Fernando Noel Dulout". Universidad Nacional de La Plata. Facultad de Ciencias Veterinarias. Instituto de Genética Veterinaria; ArgentinaFil: Barbisan, Gisela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico CONICET- La Plata. Instituto de Genética Veterinaria "Ing. Fernando Noel Dulout". Universidad Nacional de La Plata. Facultad de Ciencias Veterinarias. Instituto de Genética Veterinaria; ArgentinaFil: Poleri, Lucía Belén. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico CONICET- La Plata. Instituto de Genética Veterinaria "Ing. Fernando Noel Dulout". Universidad Nacional de La Plata. Facultad de Ciencias Veterinarias. Instituto de Genética Veterinaria; ArgentinaFil: Golijow, Carlos Daniel. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico CONICET- La Plata. Instituto de Genética Veterinaria "Ing. Fernando Noel Dulout". Universidad Nacional de La Plata. Facultad de Ciencias Veterinarias. Instituto de Genética Veterinaria; Argentin

A Kinome-wide screen identifies a CDKL5-SOX9 regulatory axis in epithelial cell death and kidney injury

© 2020, The Author(s). Renal tubular epithelial cells (RTECs) perform the essential function of maintaining the constancy of body fluid composition and volume. Toxic, inflammatory, or hypoxic-insults to RTECs can cause systemic fluid imbalance, electrolyte abnormalities and metabolic waste accumulation- manifesting as acute kidney injury (AKI), a common disorder associated with adverse long-term sequelae and high mortality. Here we report the results of a kinome-wide RNAi screen for cellular pathways involved in AKI-associated RTEC-dysfunction and cell death. Our screen and validation studies reveal an essential role of Cdkl5-kinase in RTEC cell death. In mouse models, genetic or pharmacological Cdkl5 inhibition mitigates nephrotoxic and ischemia-associated AKI. We propose that Cdkl5 is a stress-responsive kinase that promotes renal injury in part through phosphorylation-dependent suppression of pro-survival transcription regulator Sox9. These findings reveal a surprising non-neuronal function of Cdkl5, identify a pathogenic Cdkl5-Sox9 axis in epithelial cell-death, and support CDKL5 antagonism as a therapeutic approach for AKI

Human Papillomaviruses Activate the ATM DNA Damage Pathway for Viral Genome Amplification upon Differentiation

Human papillomaviruses (HPV) are the causative agents of cervical cancers. The infectious HPV life cycle is closely linked to the differentiation state of the host epithelia, with viral genome amplification, late gene expression and virion production restricted to suprabasal cells. The E6 and E7 proteins provide an environment conducive to DNA synthesis upon differentiation, but little is known concerning the mechanisms that regulate productive viral genome amplification. Using keratinocytes that stably maintain HPV-31 episomes, and chemical inhibitors, we demonstrate that viral proteins activate the ATM DNA damage response in differentiating cells, as indicated by phosphorylation of CHK2, BRCA1 and NBS1. This activation is necessary for viral genome amplification, as well as for formation of viral replication foci. In contrast, inhibition of ATM kinase activity in undifferentiated keratinocytes had no effect on the stable maintenance of viral genomes. Previous studies have shown that HPVs induce low levels of caspase 3/7 activation upon differentiation and that this is important for cleavage of the E1 replication protein and genome amplification. Our studies demonstrate that caspase cleavage is induced upon differentiation of HPV positive cells through the action of the DNA damage protein kinase CHK2, which may be activated as a result of E7 binding to the ATM kinase. These findings identify a major regulatory mechanism responsible for productive HPV replication in differentiating cells. Our results have potential implications for the development of anti-viral therapies to treat HPV infections

Changes in intracellular energy transfer enzymes in muscles of mice with deleted wolframin (wfs1) gene

Introduction To study the mechanisms of Wolfram syndrome, we assessed the changes in activities, amounts and functional coupling between mitochondria and enzymes involved in the transport of energy in muscles of wfs1‐deficient mice, models of this syndrome. Materials and methods Samples of heart, m. soleus and m. rectus femoris of wfs1‐deficient and wild‐type mice. Real‐time PCR method, spectrophotometry and nano‐LC‐MS/MS analysis of homogenates. Coupling between mitochondria and enzymes was assayed by oxygraphy of permeabilized muscle fibres. Results Compared with wild type, in m. rectus femoris of wfs1‐deficient mice mRNA level of muscle‐type creatine kinase isoform was two times (P<0.05) lower, total activities of creatine and adenylate kinase decreased by 34% (P<0.01) and 48% (P<0.02) respectively. In wfs1‐deficient mice functional coupling of adenylate kinase and mitochondria in heart decreased by 39% (P<0.05), but in m. soleus and m. rectus femoris did not change. Coupling of mitochondria and creatine kinase did not alter in any muscle of wfs1‐deficient mice. The amounts of mitochondrial sarcomeric creatine kinase in wfs1‐deficient m. rectus femoris increased 3.16‐fold (P<0.001) and mitochondrial adenylate kinase 2 2.09‐fold (P<0.01), cytoplasmatic adenylate kinase 1 but 2.12‐fold (P<0.01) decreased compared with wild‐type muscle. Conclusion In hearts of wfs1‐deficient mice, functional coupling of adenylate kinase and mitochondria decreased. Despite the drop in total activities of creatine and adenylate kinase in m. rectus femoris, the functional coupling of mitochondria did not change, because the amounts of mitochondrial isoforms of these enzymes even increased

Increased proton leak and expression of mitochondria! proteins in white skeletal muscle of mice with deleted wolframin (wfs1) gene

Poster abstrac

Expression of mitochondrial proteins and respiration in muscles of mice with deleted wolframin (wfs1) gene

Introduction Mice models of Wfs1 deficiency allow to study the mechanisms of human Wolfram syndrome. Our aim was to assess the changes in mitochondrial proteins and function in muscles of wfs1‐deficient mice. Materials and methods Samples of heart, m. soleus and m. rectus femoris of wfs1‐deficient and wild‐type mice. LFQ intensity of proteins was evaluated by nano‐LC‐MS/MS analysis of muscle homogenates, mRNA levels of uncoupler proteins, UCP2 and UCP3, by real‐time PCR method, and citrate synthase activity was determined spectrophotometrically. Mitochondrial function was assayed by high‐resolution oxygraphy of permeabilized muscle fibres. Results Compared with controls, there was no change in proton leak and citrate synthase activity in the wfs1‐deficient heart and m. soleus, but in m. rectus femoris 16.1‐fold (P<0.002) and 1.46‐fold (P<0.05) increases were found respectively. However, oxidative phosphorylation did not change in any muscle group. UCP2 mRNA level was in wfs1‐deficient m. rectus femoris 2.6 times (P<0.05) higher than in wild‐type muscle. UCP3 expression was one order of magnitude lesser than UCP2 and did not differ in muscle groups. The amounts of mitochondrial pyruvate dehydrogenase E1 component subunit alpha and mitochondrial succinate dehydrogenase iron sulphur subunit were in m. rectus femoris, compared with the wild type, respectively, 1.69 (P<0.005) and 2.08 times (P<0.0002) larger. Conclusion Wfs1‐deficient mice are characterized by a larger amount and activity of mitochondrial proteins in m. rectus femoris; the proton leak in this muscle was increased probably due to a larger amount of UCP2

Lack of dystrophin is associated with altered integration of the mitochondria and ATPases in slow-twitch muscle cells of MDX mice

Braun U, Paju K, Eimre M, et al. Lack of dystrophin is associated with altered integration of the mitochondria and ATPases in slow-twitch muscle cells of MDX mice. BIOCHIMICA ET BIOPHYSICA ACTA-BIOENERGETICS. 2001;1505(2-3):258-270.The potential role of dystrophin-mediated control of systems integrating mitochondria with ATPases was assessed in muscle cells. Mitochondrial distribution and function in skinned cardiac and skeletal muscle fibers from dystrophin-deficient (MDX) and wild-type mice were compared. Laser confocal microscopy revealed disorganized mitochondrial arrays in m. gastrocnemius in MDX mice, whereas the other muscles appeared normal in this group. Irrespective of muscle type, the absence of dystrophin had no effect on the maximal capacity of oxidative phosphorylation, nor on coupling between oxidation and phosphorylation. However, in the myocardium and m. soleus, the coupling of mitochondrial creatine kinase to adenine nucleotide translocase was attenuated as evidenced by the decreased effect of creatine on the K-m for ADP in the reactions of oxidative phosphorylation. In m. soleus, a low K-m, for ADP compared to the wild-type counterpart was found, which implies increased permeability for that nucleotide across the mitochondrial outer membrane. In normal cardiac fibers 35% of the ADP flux generated by ATPases was not accessible to the external pyruvate kinase-phosphoenolpyruvate system, which suggests the compartmentalized (direct) channeling of that fraction of ADP to mitochondria. Compared to control, the direct ADP transfer was increased in MDX ventricles. In conclusion. our data indicate that in slow-twitch muscle cells, the absence of dystrophin is associated with the rearrangement of the intracellular energy and feedback signal transfer systems between mitochondria and ATPases. As the mechanisms mediated by creatine kinases become ineffective, the role of diffusion of adenine nucleotides increases due to the higher permeability of the mitochondrial outer membrane for ADP and enhanced compartmentalization of ADP flux. (C) 2001 Elsevier Science B.V. All rights reserved