No description available.

L’anémie de Fanconi (FANC) est une maladie génétique humaine causant chez les patients une anémie sévère, une susceptibilité accrue à certains cancers, ainsi que des anomalies du développement dont un hypogonadisme. La voie FANC est impliquée dans la réponse au stress réplicatif et la réparation de l’ADN, et joue un rôle potentiel dans la régulation physiologique des cellules souches fœtales et adultes. Dans les modèles murins de la voie FANC, le phénotype majeur est une infertilité liée à un problème de développement du lignage germinal, qui est un paradigme pour l’étude des mécanismes contribuant à la pathologie. Notre étude a porté sur la caractérisation du défaut des cellules germinales primordiales (CGP) dans les embryons murins fancg-/-. Nous avons mis en évidence un défaut numérique des CGP très tôt dans le développement du lignage germinal dès les stades 9,5-10,5 jours post coïtum (jpc). Les CGP présentent un léger défaut de la prolifération à 10,5-11,5 jpc, mais aucun blocage de cycle. L’atteinte proliférative semble trop faible pour expliquer à elle seule la diminution drastique du nombre de CGP, ainsi que le profil mosaïque des gonades embryonnaires avec la présence de cordons dépourvus de CGP observé à 13,5 jpc. L’étude in vivo et ex vivo du comportement migratoire des CGP fancg-/- a permis de mettre en évidence des anomalies de la motilité des CGP, probablement liée à une activation anormale de la GTPase RAC1. Nous avons observé à 11,5 jpc une diminution du nombre de cellules au niveau du front de migration de la population de CGP. Ces anomalies entraîneraient ainsi un retard de migration et l’incapacité pour une fraction des GCP fancg-/- d’atteindre correctement les crêtes génitales à 11,5 jpc. La déplétion des CGP semble ainsi liée principalement à un défaut migratoire conduisant à une augmentation de la mort des CGP à 11,5 jpc, associé au défaut prolifératif intrinsèque des CPG fancg-/-. / Fanconi Anemia (FANC) is a genetic human disease, causing in patient a severe anemia, a higher risk to develop some cancers, and developmental anomalies including hypogonadism. The FANC pathway is involved in the replicative stress response and DNA repair, and has a potential role in the physiological regulation of fetal and adult stem cells. In mice models of the FANC pathway, the main phenotype is the sterility issue, associated to a developmental defect of the germ cell lineage, and is so a paradigm to study the pathology. Our study aimed to characterize the primordial germ cell (PGC) defect in fancg-/- mouse embryos. We showed a numerical defect of PGC early in the germ cell lineage development, as soon as 9.5–10.5 days post coitum (dpc). PGC show a mild defect of proliferation at 10.5–11.5 dpc, but no cell cycle arrest. This low proliferative defect can neither fully explain the drastic decrease of the PGC number, nor the mosaic profile of fetal gonads displaying cords without PGC at 13.5 dpc. In vitro and ex vivo studies of the migration behavior of fancg-/- PGC highlight abnormalities of the PGC’s motility, probably linked to abberant RAC1 GTPase activity. We also observed at 11.5 dpc a decrease of the cell number at the front of migration of the PGC population in 11.5 dpc fancg-/- embryos. Those motility defects could induce a migration delay, preventing a fraction of the fancg-/- PGC population to reach correctly the genital crests by 11.5 dpc. Hence, PGC depletion seems mainly linked to a migratory defect leading to increased cell death in PGC at 11.5 dpc, associated to an intrinsic proliferation defect of fancg-/- PGC.

京都大学 / 0048 / 新制・課程博士 / 博士(農学) / 甲第17793号 / 農博第2014号 / 新制||農||1016(附属図書館) / 学位論文||H25||N4784(農学部図書室) / 30600 / 京都大学大学院農学研究科応用生物科学専攻 / (主査)教授 今井 裕, 教授 久米 新一, 教授 松井 徹 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DFAM

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第18172号 / 医博第3892号 / 新制||医||1003(附属図書館) / 31030 / 京都大学大学院医学研究科医学専攻 / (主査)教授 篠原 隆司, 教授 中辻 憲夫, 教授 萩原 正敏, 教授 小西 郁生 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DGAM

京都大学 / 0048 / 新制・課程博士 / 博士(医科学) / 甲第20285号 / 医科博第76号 / 新制||医科||5(附属図書館) / 京都大学大学院医学研究科医科学専攻 / (主査)教授 篠原 隆司, 教授 浅野 雅秀, 教授 近藤 玄 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

京都大学 / 0048 / 新制・課程博士 / 博士(農学) / 甲第21166号 / 農博第2292号 / 新制||農||1061(附属図書館) / 学位論文||H30||N5140(農学部図書室) / 京都大学大学院農学研究科応用生物科学専攻 / (主査)教授 今井 裕, 教授 久米 新一, 准教授 南 直治郎 / 学位規則第4条第1項該当 / Doctor of Agricultural Science / Kyoto University / DFAM

No description available.

No description available.

Assessment of chemical mutagenicity is essential to protecting human health from genetic disease. Current assays are limited in their ability to provide mechanistic insight into the endogenous and exogenous processes involved in mutagenesis. Duplex Sequencing (DS), a novel error-corrected sequencing technology, overcomes many of the limitations faced by conventional mutagenicity assays. DS could be used to eliminate reliance on standalone reporter assays and provide mechanistic information alongside mutation frequency (MF) data. Furthermore, customizable panels enable assessment of the endogenous genomic features that drive mutagenesis. However, the performance of DS must be thoroughly assessed before it can be routinely implemented for standard testing. The objectives of this study were to demonstrate the potential of DS as a robust in vivo mutagenicity test and to explore its rich data to gain a better understanding of spontaneous and chemically-induced mutagenicity in somatic and germ cells. We used DS to study spontaneous and procarbazine (PRC)-induced mutations in the bone marrow (BM) and germ cells of MutaMouse males across a panel of 20 diverse genomic targets. Mice were exposed to 0, 6.25, 12.5, or 25 mg/kg-bw/day for 28 days by oral gavage and tissues were sampled at least 28 days post-exposure. Results were compared with those obtained using the conventional lacZ viral plaque assay on the same samples. DS detected significant increases in MF and distinct spectra consistent with the known mutagenic mechanisms of PRC in both tissues. Mouse PRC doses at which significant effects were observed are in range with those used for chemotherapy, suggesting that similar effects may be observed in human patients. This supports the contribution of PRC towards secondary cancers following treatment. DS results were comparable to those obtained using the gold-standard lacZ TGR assay, with DS showing greater sensitivity to detect smaller changes in MF. Analysis of mutation spectra and the genomic features that drive the mutational response revealed intrinsic differences between BM and germ cells that may underlie differences in endogenous mutagenic mechanisms and/or DNA repair pathways. The results suggest that germ cells may have intrinsic mechanisms to reduce mutation burden relative to somatic cells. While historically analysis of germ cell mutagenicity has been neglected in favour of somatic cells, our work supports the independent assessment of germ cell mutagenicity during regulatory testing. Finally, we conducted power analyses to inform the optimal DS study designs for the two tissues. We found that low intra-group variability within BM samples allows a reduction in sample size to three animals per group whilst still maintaining 80% power to detect an effect. In contrast, the relatively high intra-group variability and low background MF in germ cells suggests a minimum of eight animals per group to detect an effect. Overall, our results support the use of DS as a mutagenicity test and highlight many of the advantages it holds over conventional assays. Moreover, our study reveals the potential for mutagenic effects in PRC-treated cancer patients. Further work to test DS with more chemicals and across a wider range of tissues is recommended for future implementation as a mutagenicity test.

The genetic variation among extant life forms reflects the outcomes of evolution. The fodder of evolution – germline mutations – is shaped by the interplay among evolutionary forces – notably natural selection and random genetic drift. In turn, these forces leave footprints recorded in the genetic variation of extant life forms. Characterizing these footprints to understand how evolution works is at the heart of population genetics. To this end, massive datasets of genetic variation have opened new avenues of research, around how mutation rates evolve for instance, and reinvigorated long standing questions in population genetics, notably about the genetic basis of adaptation. In turn, theoretical models of evolution inform what kind of footprints we expect evolution to leave behind in such data. Two theoretical models that investigate open questions in population genetics are described in this thesis. In Chapter 1, I consider the evolution of germline mutation rates, particularly on short evolutionary timescales, and ask if recently observed variation in mutation rates among human lineages could be explained by evolution at genetic modifiers of mutation rates. Genetic modifiers of mutation rates are expected to evolve under purifying selection: mutations at modifiers that increase mutation rates (“mutator alleles”) should be selected against, because they increase the burden of deleterious mutations in individuals who carry them. The frequencies of mutator alleles are also affected by mutation, genetic drift, and demographic processes. We model the evolution of mutator alleles under the interplay of these forces and characterize the dynamics at mutation rate modifiers as a function of the efficacy of selection acting on them. We find that modifiers under intermediate selection have the greatest contribution to variation in mutation rates between distantly related populations, but only variation at strongly selected modifiers turns over fast enough to explain variation in mutation rates among human lineages. We also predict that strongly selected modifiers could be potentially identified in the contemporary datasets of human pedigrees used to study germline mutations. In Chapter 2, I consider a central and enduring question in evolutionary biology: whether adaptation typically arises from few large effect changes or from many small effect changes. Both sides are supported by ample evidence. Yet it is unclear how to translate this evidence into general answers about the genetic basis of adaptation, in part because different methodologies have different limitations and ask different questions. Theory may offer a way out of this quagmire or at least a start. To this end, we reframe the question in terms of traits and ask: how does the genetic basis of adaptation depend on the ecological and genetic attributes of a trait? To start answering this question, I model adaptation in a simple yet highly relevant setting. I consider a trait under stabilizing selection and assume the distribution of trait values in the population is initially at mutation-selection-drift-balance. I then characterize the adaptive response that is elicited by a sudden change in the environment. I find that the adaptive response, and notably the probability that adaptation arises from the fixation of large effect alleles, depends on the size of the environmental change and the genetic architecture of the trait. These attributes are measurable and can be directly related to the disparate evidence that we have about the genetic basis of adaptation. Thus, this kind of modeling may help translate such evidence into general conclusions about how traits evolve. My thesis work was interrupted by the global COVID-19 pandemic, and in response to this pandemic, governments around the world implemented shelter-in-place protocols. However, essential workers were exempt from these protocols, potentially decreasing their efficacy. In Chapter 3, we describe our epidemiological project, aimed at understanding the impact of essential workers on epidemic control. To this end, we model three different archetypes of essential workers under a reasonably realistic SEIR model of the COVID-19 pandemic. We find that the different social interactions that essential workers maintain qualitatively changes their personal risk of infection and the spread of the overall epidemic. These results highlight the utility of not considering essential workers as a monolithic group but instead distinguishing between the impact of different types of essential workers on epidemic control.