Orientador: Roberto André Kraenkel / Banca: Hilda Cerdeira / Banca: André Nachbin / Resumo: Neste trabalho estudamos uma equação diferencial com retardo, a equação de Hutchinson, que é um modelo simples para a dinâmica de uma população que exibe dependência em tempos passados por meio de uma variável defasada. Essa equação pode ser vista como uma equação mínima que é capaz de dar origem a soluções oscilatórias em modelos de uma única espécie. Para analisar a solução próxima do ponto de bifurcação em que surgem as oscilações, empregamos o método de múltiplas escalas. Os resultados obtidos mostram as próprias limitações do método, concordando apenas parcialmente com os resultados numéricos. Também analisamos uma variante da equação de Hutchinson com capacidade de suporte dependente do tempo e periódica, e vimos que, mesmo com amplitudes de perturbação muito pequenas, o acoplamento entre as frequências da capacidade de suporte e da oscilação natural pode ter um efeito pronunciado sobre a dinâmica da população. Apresentamos uma análise de ressonâncias para este caso e mostramos a existência de frequências da capacidade de suporte perto das quais a solução da equação exibe um comportamento inesperado / Abstract: In the present work, we study a delay diUerential equation, namely the Hutchinson equation, which is a simple model for the dynamics of a population that shows dependence on past times through a lagged variable. This equation can be seen as a minimal equation that is able to produce oscillatory solutions in single species models. In order to analyze the solution near the bifurcation point at which oscillations set in, we employ the multiple scales method. The results obtained expose the limitations of the method, agreeing only partially with numerical results. We also analyze a variation of the Hutchinson equation with a periodic timevarying carrying capacity, and Vnd that, even at very small amplitude perturbations, the coupling of frequencies between the carrying capacity oscillation and the natural oscillation can have a major eUect on the population dynamics. We present a resonance analysis for this case and show the existence of carrying capacity frequencies near which the equation's solution exhibit an unexpected behavior / Mestre

The discovery of mammalian protein prenylation was originally motivated by an effort to identify a nonsterol isoprenoid which indirect evidence suggested was a coregulator of isoprenoid biosynthesis and played a critical role in cellular proliferation. The first prenylated proteins to be identified were the nuclear lamin proteins-B lamins and prelamin A-which were subsequently shown to be farnesylated at a carboxyl-terminal CAAX motif. In both types of lamin, the farnesylation and carboxymethylation play a role in targeting these proteins to the nuclear envelope. The nucleus can be demonstrated to be a CAAX processing compartment for the lamins. In the case of prelamin A, there is removal of a carboxyl-terminal polypeptide which is specifically catalyzed by the enzyme Zmpste24. This processing event is necessary for assembly of lamin A into the lamina and may play a role in cell cycle control. Because the nucleus contains only one target membrane, lamin farnesylation and carboxymethylation may be sufficient to allow association with this membrane. This stands in contrast to farnesylated proteins expressed in the cytoplasm.

Algumas síndromes genéticas monogênicas apresentam fenótipos considerados progeróides, ou seja, desenvolvem precocemente características clínicas semelhantes às observadas no envelhecimento humano normal. A relação fisiopatológica entre essas doenças e o processo de envelhecimento vem sendo estudada, sendo que o entendimento dos mecanismos moleculares em um campo contribui para a compreensão do outro. As síndromes de Hutchinson-Gilford e Rothmund-Thomson são duas condições progeróides raras, já bem caracterizadas clinicamente, que são causadas por alterações nos genes LMNA (em um alelo) e RECQL4 (nos dois alelos), respectivamente. No entanto, em cerca de 40% a 60% dos indivíduos com a síndrome de Rothmund-Thomson, não são encontradas mutações em RECQL4, constituindo um subgrupo chamado de tipo I; desse modo, os casos com alteração no gene constituem o tipo II da síndrome. Indivíduos com o tipo II apresentam um risco aumentado para o desenvolvimento de câncer, particularmente o osteossarcoma. Neste trabalho, nove pacientes com sinais progeróides foram avaliados clinicamente e tiveram o DNA sequenciado. Um paciente recebeu o diagnóstico clínico de síndrome de Hutchinson-Gilford, que foi confirmado pela mutação patogênica mais frequente encontrada no gene LMNA (p.Gly608Gly). Oito pacientes jovens, com mediana de idade de 2 anos e 2 meses, foram diagnosticados clinicamente como afetados pela síndrome de Rothmund-Thomson, cujas características clínicas mais comuns incluíam déficit pôndero-estatural, cabelos e sobrancelhas/cílios esparsos, fronte ampla, lesão cutânea (eritema em face e lesão poiquilodérmica) e anomalias ósseas, alterações típicas da síndrome. Catarata estava presente em 50% dos indivíduos. Nenhum dos pacientes desenvolveu algum tipo de tumor até o momento. O sequenciamento do gene RECQL4 mostrou a presença de três variantes patogênicas diferentes, em três probandos (37,5%), sendo dois em homozigose e um em heterozigose composta, todas já descritas previamente na literatura. Em busca de alterações em outro gene que pudesse explicar o quadro apresentado pelos pacientes sem mutação, três dos probandos, incluindo os genitores de um deles, tiveram o exoma sequenciado. No entanto, não foram encontradas, nessa etapa, variantes adicionais que explicassem na totalidade os fenótipos apresentados. Comparando os achados clínicos dos pacientes com a síndrome de Rothmund-Thomson tipo I e tipo II, foi observada diferença estatisticamente significante na incidência de catarata subcapsular nos pacientes sem mutação (p<0,05), semelhante ao que é descrito na literatura. Diante dos achados clínicos e moleculares obtidos, foi realizado o aconselhamento genético para todos os indivíduos, enfatizando a evolução, os cuidados e acompanhamentos necessários para as duas doenças em questão e fornecendo informações aos genitores dos probandos sobre o risco de recorrência para a prole futura / Some monogenic disorders exhibit progeroid phenotypes, in other words, they develop premature characteristics similar to those observed in normal human aging. The physiopathological correlation between these diseases and the aging process has being studied, and the understanding of the molecular mechanisms in one field contributes to the understanding of the other. The Hutchinson-Gilford and Rothmund-Thomson syndromes are two clinically well-characterized rare progerioid conditions that are caused by changes in the LMNA (in one allele) and RECQL4 (in both alleles) genes, respectively. However, in about 40% to 60% of individuals with Rothmund-Thomson syndrome, no mutations are found in RECQL4, constituting a subgroup called type I; thus, cases with mutations in the gene constitute the type II group of the syndrome. Individuals with type II have an increased risk for the development of cancer, particularly osteosarcoma. In this study, nine patients with progerioid signs were clinically evaluated and had the DNA sequenced. One patient was clinically diagnosed with Hutchinson-Gilford syndrome, which was confirmed by the most frequent pathogenic mutation found in the LMNA gene (p.Gly608Gly). Eight young patients with median age of 2 years and 2 months were clinically diagnosed as affected by Rothmund-Thomson syndrome, whose most common clinical features included: short stature; sparse hair, eyebrows and eyelashes; erythematous skin lesions and poikiloderma; and bone abnormalities; all typical of the syndrome. Cataract was present in 50% of individuals. None of the patients has developed any type of tumor at this time. Sequencing of the RECQL4 gene showed the presence of three different pathogenic variants in three probands (37.5%), two in homozygous and one in compound heterozygosity, all previously described in the literature. In search of alterations in another gene that could explain the phenotype presented by the patients without mutation, three of the probands, including the parents of one of them, had the exoma sequenced. However, there were no additional variants at this stage that fully explained the phenotypes presented by these individuals. Comparing the clinical findings of patients with Rothmund-Thomson syndrome type I and type II, a statistically significant difference was observed in the incidence of subcapsular cataract in patients without mutation (p <0.05), similar to that described in the literature. In light of the clinical and molecular findings, genetic counseling was performed for all individuals, emphasizing the evolution, care and follow-up needed for the two diseases in question and providing information to the parents of the probands on the risk of recurrence for future offspring

Algumas síndromes genéticas monogênicas apresentam fenótipos considerados progeróides, ou seja, desenvolvem precocemente características clínicas semelhantes às observadas no envelhecimento humano normal. A relação fisiopatológica entre essas doenças e o processo de envelhecimento vem sendo estudada, sendo que o entendimento dos mecanismos moleculares em um campo contribui para a compreensão do outro. As síndromes de Hutchinson-Gilford e Rothmund-Thomson são duas condições progeróides raras, já bem caracterizadas clinicamente, que são causadas por alterações nos genes LMNA (em um alelo) e RECQL4 (nos dois alelos), respectivamente. No entanto, em cerca de 40% a 60% dos indivíduos com a síndrome de Rothmund-Thomson, não são encontradas mutações em RECQL4, constituindo um subgrupo chamado de tipo I; desse modo, os casos com alteração no gene constituem o tipo II da síndrome. Indivíduos com o tipo II apresentam um risco aumentado para o desenvolvimento de câncer, particularmente o osteossarcoma. Neste trabalho, nove pacientes com sinais progeróides foram avaliados clinicamente e tiveram o DNA sequenciado. Um paciente recebeu o diagnóstico clínico de síndrome de Hutchinson-Gilford, que foi confirmado pela mutação patogênica mais frequente encontrada no gene LMNA (p.Gly608Gly). Oito pacientes jovens, com mediana de idade de 2 anos e 2 meses, foram diagnosticados clinicamente como afetados pela síndrome de Rothmund-Thomson, cujas características clínicas mais comuns incluíam déficit pôndero-estatural, cabelos e sobrancelhas/cílios esparsos, fronte ampla, lesão cutânea (eritema em face e lesão poiquilodérmica) e anomalias ósseas, alterações típicas da síndrome. Catarata estava presente em 50% dos indivíduos. Nenhum dos pacientes desenvolveu algum tipo de tumor até o momento. O sequenciamento do gene RECQL4 mostrou a presença de três variantes patogênicas diferentes, em três probandos (37,5%), sendo dois em homozigose e um em heterozigose composta, todas já descritas previamente na literatura. Em busca de alterações em outro gene que pudesse explicar o quadro apresentado pelos pacientes sem mutação, três dos probandos, incluindo os genitores de um deles, tiveram o exoma sequenciado. No entanto, não foram encontradas, nessa etapa, variantes adicionais que explicassem na totalidade os fenótipos apresentados. Comparando os achados clínicos dos pacientes com a síndrome de Rothmund-Thomson tipo I e tipo II, foi observada diferença estatisticamente significante na incidência de catarata subcapsular nos pacientes sem mutação (p<0,05), semelhante ao que é descrito na literatura. Diante dos achados clínicos e moleculares obtidos, foi realizado o aconselhamento genético para todos os indivíduos, enfatizando a evolução, os cuidados e acompanhamentos necessários para as duas doenças em questão e fornecendo informações aos genitores dos probandos sobre o risco de recorrência para a prole futura / Some monogenic disorders exhibit progeroid phenotypes, in other words, they develop premature characteristics similar to those observed in normal human aging. The physiopathological correlation between these diseases and the aging process has being studied, and the understanding of the molecular mechanisms in one field contributes to the understanding of the other. The Hutchinson-Gilford and Rothmund-Thomson syndromes are two clinically well-characterized rare progerioid conditions that are caused by changes in the LMNA (in one allele) and RECQL4 (in both alleles) genes, respectively. However, in about 40% to 60% of individuals with Rothmund-Thomson syndrome, no mutations are found in RECQL4, constituting a subgroup called type I; thus, cases with mutations in the gene constitute the type II group of the syndrome. Individuals with type II have an increased risk for the development of cancer, particularly osteosarcoma. In this study, nine patients with progerioid signs were clinically evaluated and had the DNA sequenced. One patient was clinically diagnosed with Hutchinson-Gilford syndrome, which was confirmed by the most frequent pathogenic mutation found in the LMNA gene (p.Gly608Gly). Eight young patients with median age of 2 years and 2 months were clinically diagnosed as affected by Rothmund-Thomson syndrome, whose most common clinical features included: short stature; sparse hair, eyebrows and eyelashes; erythematous skin lesions and poikiloderma; and bone abnormalities; all typical of the syndrome. Cataract was present in 50% of individuals. None of the patients has developed any type of tumor at this time. Sequencing of the RECQL4 gene showed the presence of three different pathogenic variants in three probands (37.5%), two in homozygous and one in compound heterozygosity, all previously described in the literature. In search of alterations in another gene that could explain the phenotype presented by the patients without mutation, three of the probands, including the parents of one of them, had the exoma sequenced. However, there were no additional variants at this stage that fully explained the phenotypes presented by these individuals. Comparing the clinical findings of patients with Rothmund-Thomson syndrome type I and type II, a statistically significant difference was observed in the incidence of subcapsular cataract in patients without mutation (p <0.05), similar to that described in the literature. In light of the clinical and molecular findings, genetic counseling was performed for all individuals, emphasizing the evolution, care and follow-up needed for the two diseases in question and providing information to the parents of the probands on the risk of recurrence for future offspring

Le vieillissement est un processus complexe qui peut être influencé par des facteurs environnementaux et génétiques. Le syndrome progéroïde de Hutchinson-Gilford (HGPS ou progéria) fourni une preuve irréfutable de l'implication de l'épissage dans le processus de vieillissement. La progéria est une maladie due à une mutation hétérozygote silencieuse qui renforce l'utilisation d'un site 5' d'épissage interne dans l'exon 11 de l'ARN pré-messager LMNA, ce qui entraîne la production d'une protéine tronquée appelée «progérine». Le défaut d'épissage du gène LMNA a aussi lieu dans les cellules de personnes âgées, et la correction de ce défaut permet un sauvetage partiel des anomalies qu'il provoque. Ceci fait de l'ARN pré-messager LMNA une cible très attractive pour des thérapies ayant pour but de corriger l'épissage. Mes travaux de thèse ont montré que cette mutation silencieuse active un site d'épissage 5' dans l'exon 11 en changeant la structure de l'ARN. Ce changement de structure facilite l'interaction de la snRNP U1 avec le site d'épissage et permet ainsi sa modulation par les protéines SR SRSF1 et SRSF6. J'ai aussi participé à la caractérisation d'un nouveau modèle murin qui reproduit l'altération d'épissage des patients HGPS au niveau du gène Lmna souris. De façon surprenante, ce modèle récapitule tous les phénotypes du syndrome HGPS. Les souris homozygotes, dans lesquelles la plupart de la lamine A est convertie en progérine, ne vivent pas plus de 5 mois, alors que les souris hétérozygotes vivent autour d'un an et que les contrôles sauvages vivent deux ans. Étonnamment, des souris qui n'expriment ni la lamine A ni la progérine, mais uniquement de la lamine C, vivent plus longtemps que les souris contrôle, suggérant que la lamine A et la progérine, qui sont produites à partir du même transcrit, participent à la régulation de la durée de la vie. De plus, la caractérisation initiale des souris HGPS indique que l'expression de la progérine est délétère pour le tissu adipeux, établissant ainsi un lien inattendu entre l'épuisement du tissu adipeux et le vieillissement accéléré. Ce nouveau modèle murin est actuellement en train d'être utilisé pour des approches de modulation de l'épissage aberrant du gène LMNA avec des oligonucléotides antisense et des petites molécules chimiques. / Aging is a complex cellular and organismal process that can be influenced by environmental as well as genetic factors. A striking proof-of-concept that splicing regulation plays an important role in the aging process is provided by Hutchinson-Gilford progeria syndrome (HGPS), a disease caused by a heterozygous silent mutation that enhances the use of an internal 5' splice site in exon 11 of LMNA pre-mRNA and leads to the production of a truncated protein called “progerin”. The LMNA splicing defect also occurs with increased frequency in cells from healthy aged individuals and correction of this defect leads to partial reversal of age-related dysfunction. This makes LMNA pre-mRNA an attractive target for splicing-correction therapies. During my PhD thesis I have characterized the splicing mechanism responsible for progerin production and demonstrated that this process is conserved from mouse to human. I have found that HGPS mutation changes the accessibility of the exon 11 internal 5' splice site, allowing its modulation by U1 snRNP and a subset of SR proteins, namely SRSF6 and SRSF1. I have also participated to the characterization of a new mouse model reproducing human HGPS splicing alteration in the mouse Lmna gene. Strikingly, this model recapitulates all phenotypic manifestations of HGPS. The homozygous mice, where most lamin A is converted to progerin, lived no longer than 5 months, whereas heterozygous mice lived in average one year and wild type littermates up to two years. Unexpectedly, mice expressing neither lamin A nor progerin, but only lamin C, lived longer than wild type littermates mice, suggesting that lamin A and progerin which are produced from the same transcript, control critical steps of lifespan. Furthermore, initial characterization of HGPS mouse model indicated that progerin expression is deleterious for adipose tissue, establishing an unexpected link between adipose tissue depletion and accelerated aging. The new mouse model is currently being used for pharmacological modulation of LMNA aberrant splicing by antisense oligonucleotides and small molecules.

Call number: LD2668 .T4 1965 L42 / Master of Science

Thesis (D. Min.)--New Orleans Baptist Theological Seminary, 1999. / Abstract and vita. Includes bibliographical references (leaves 104-107).

Thesis (Ph.D.)--Baylor University, 2008. / Includes bibliographical references (p. 147-152)

Thesis (D. Min.)--New Orleans Baptist Theological Seminary, 1999. / Abstract and vita. Includes bibliographical references (leaves 104-107).

Thesis (D. Min.)--New Orleans Baptist Theological Seminary, 1999. / Abstract and vita. Includes bibliographical references (leaves 104-107).