Equine sarcoids are the most common skin tumors of horses. Despite being such a common entity, relatively little is known about many features of sarcoid epidemiology or growth. In addition, due to the detection of Bovine Papillomavirus (BPV) DNA of 2 different types, BPV type 1 (BPV1) and BPV type 2 (BPV2), in equine sarcoids BPV has been suggested as the causative agent of sarcoid development. Recently, however, BPV DNA has also been detected in other skin conditions of horses; the significance of this is unclear. Multiple studies to learn more about sarcoids were undertaken.<p> To investigate the epidemiology of sarcoids in horses in Western Canada the records of five veterinary diagnostic laboratories were searched to identify submissions of sarcoids from horses. The submission record and diagnostic reports of 802 separate submissions of equine sarcoids were reviewed for age, breed, and gender of the horse and the number, location and clinical type of sarcoid. Based on these submissions, horses of a wide variety of ages and 23 different equine breeds were affected, within these breeds, Donkeys were overrepresented.<p> The presence of BPV was determined by Polymerase Chain Reaction (PCR). BPV was found in 74 of 96 (77.1%) samples, and using Restriction Fragment Length Polymorphism, BPV1 and BPV2 were identified in these samples. BPV2 was present in 59 (79.7%) of these. Unlike other areas in the world, in Western Canada, equine sarcoids are most commonly associated with BPV2.<p> A second study examined different clinical types of sarcoids to determine if there was differential expression of immunohistochemical markers associated with apoptosis, Cleaved Caspase 3(ClC3), and antiapoptotic factors, B-Cell Lymphoma 2 (Bcl-2) and Survivin. No differences in the expression of any of these markers regardless of BPV type were noted. Survivin was expressed in equine sarcoids of all types and increased levels of expression are associated with more aggressive clinical behaviour.<p> Finally, the location of BPV DNA was determined in both sarcoids and a variety of non-sarcoid inflammatory skin conditions of horses, as well as, normal skin. PCR for BPV DNA was performed on 86 skin biopsies from horses with non-sarcoid skin conditions, as well as, normal skin. BPV DNA was present in 41 of 86 biopsies. These positive samples, in addition to BPV positive sarcoid samples from the earlier study, were dissected into tissue compartments using laser microdissection followed by 2 forms of BPV DNA amplification, PCR and isothermal loop mediated amplification. BPV DNA was more often located in the epidermis of non-sarcoid skin conditions than in sarcoids. In addition, areas of inflammation within the dermis and epidermis were more likely to contain BPV DNA than non-inflamed areas. These results suggest that while BPV is commonly found in equine skin, the location where it is found differs between sarcoids and non-sarcoid samples. When BPV DNA was found in non-sarcoid samples, it was commonly associated with inflammation suggesting that microscopic damage to the epidermal barrier of the skin maybe an adequate predisposing factor to the development of sarcoids.

Equine sarcoids are the most common skin tumors of horses. Despite being such a common entity, relatively little is known about many features of sarcoid epidemiology or growth. In addition, due to the detection of Bovine Papillomavirus (BPV) DNA of 2 different types, BPV type 1 (BPV1) and BPV type 2 (BPV2), in equine sarcoids BPV has been suggested as the causative agent of sarcoid development. Recently, however, BPV DNA has also been detected in other skin conditions of horses; the significance of this is unclear. Multiple studies to learn more about sarcoids were undertaken.<p> To investigate the epidemiology of sarcoids in horses in Western Canada the records of five veterinary diagnostic laboratories were searched to identify submissions of sarcoids from horses. The submission record and diagnostic reports of 802 separate submissions of equine sarcoids were reviewed for age, breed, and gender of the horse and the number, location and clinical type of sarcoid. Based on these submissions, horses of a wide variety of ages and 23 different equine breeds were affected, within these breeds, Donkeys were overrepresented.<p> The presence of BPV was determined by Polymerase Chain Reaction (PCR). BPV was found in 74 of 96 (77.1%) samples, and using Restriction Fragment Length Polymorphism, BPV1 and BPV2 were identified in these samples. BPV2 was present in 59 (79.7%) of these. Unlike other areas in the world, in Western Canada, equine sarcoids are most commonly associated with BPV2.<p> A second study examined different clinical types of sarcoids to determine if there was differential expression of immunohistochemical markers associated with apoptosis, Cleaved Caspase 3(ClC3), and antiapoptotic factors, B-Cell Lymphoma 2 (Bcl-2) and Survivin. No differences in the expression of any of these markers regardless of BPV type were noted. Survivin was expressed in equine sarcoids of all types and increased levels of expression are associated with more aggressive clinical behaviour.<p> Finally, the location of BPV DNA was determined in both sarcoids and a variety of non-sarcoid inflammatory skin conditions of horses, as well as, normal skin. PCR for BPV DNA was performed on 86 skin biopsies from horses with non-sarcoid skin conditions, as well as, normal skin. BPV DNA was present in 41 of 86 biopsies. These positive samples, in addition to BPV positive sarcoid samples from the earlier study, were dissected into tissue compartments using laser microdissection followed by 2 forms of BPV DNA amplification, PCR and isothermal loop mediated amplification. BPV DNA was more often located in the epidermis of non-sarcoid skin conditions than in sarcoids. In addition, areas of inflammation within the dermis and epidermis were more likely to contain BPV DNA than non-inflamed areas. These results suggest that while BPV is commonly found in equine skin, the location where it is found differs between sarcoids and non-sarcoid samples. When BPV DNA was found in non-sarcoid samples, it was commonly associated with inflammation suggesting that microscopic damage to the epidermal barrier of the skin maybe an adequate predisposing factor to the development of sarcoids.

Submitted by Caroline Falcao (caroline.rfalcao@ufpe.br) on 2017-04-04T18:49:12Z No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) 2011-Dissertação-ElydaLima.pdf: 2803583 bytes, checksum: 5b93b96556bc230adb5bb7ca5a4fb423 (MD5) / Made available in DSpace on 2017-04-04T18:49:12Z (GMT). No. of bitstreams: 2 license_rdf: 1232 bytes, checksum: 66e71c371cc565284e70f40736c94386 (MD5) 2011-Dissertação-ElydaLima.pdf: 2803583 bytes, checksum: 5b93b96556bc230adb5bb7ca5a4fb423 (MD5) Previous issue date: 2011-03-16 / A bovinocultura é um dos principais destaques do agronegócio brasileiro no cenário mundial. No entanto, algumas doenças vêm causando prejuízos consideráveis entre elas está a papilomatose bovina, uma doença infectocontagiosa, de caráter tumoral, com etiologia relacionada a infecção pelo papilomavírus bovino (BPV) que se caracteriza pela formação de tumores em tecidos da pele e mucosa. Hoje se conhecem 11 tipos diferentes de Papilomavírus bovino, sendo os BPvs tipos 1, 2 e 4 oncogênicos. Até o momento não existe vacinas para o controle ou tratamento das papilomaviroses. Diferentes estudos vêm demonstrando que a proteína L2 pode ser uma candidata ao desenvolvimento de estratégias vacinais profiláticas contra o BPV. Neste trabalho, tivemos como objetivo construir um vetor vacinal a partir do plasmídeo pCINeo (Promega®) e do gene L2 de BPV1, avaliando in vitro a expressão do antígeno L2 em células de mamíferos. O gene L2 foi amplificado pela técnica de PCR a partir do genoma completo de BPV1, com uso de oligonucleotídeos específicos contendo um epítopo AU1 no primer forwarde posteriormente clonado em vetor de passagem pGEM-T Easy (Promega®) e subclonado no vetor de expressão pCIneo, gerando a construção pCIL2B1. O plasmídeo foi transfectado in vitroem células 293 para análise funcional da expressão de L2. Os resultados obtidos confirmaram a construção pCIL2B1 por PCR e sequenciamento. A capacidade desta construção expressar o gene L2 e produzir a respectiva proteína em células de mamífero foi confirmada por RT-PCR e western blot (usando anticorpo contra o epítopo AU1). / The cattle industry is one of the main highlights of the Brazilian agribusiness on the international stage. However, some diseases have caused considerable damage including bovine papillomatosis which is an infectious tumorous disease related to bovine papillomavirus (BPV) etiology and characterized by the formation of tumors in tissues of the skin and mucosa. Currently we know 11 different types of bovine papillomavirus, among which the BPv types 1, 2 and 4 are oncogenic. So far there is no vaccine or treatment for the papilomaviroses control. Different studies have reported that the L2 protein may be a candidate for the development of prophylactic vaccine strategies against BPV. The L2 protein has a potential cross-reaction with different BPVs and HPV (HumanPapillomavirus). On this paper, our objective was to construct a vector vaccine from pCINeo plasmid (Promega ®) and the gene of BPV1 L2, evaluating in vitro L2 antigen expression in mammalian cells.The L2 gene was amplifiedby PCR from thecomplete genome ofBPV1, using specifico ligo nucleotidescontainingan AU1epitopein the for wardprimerand subsequently cloned intop GEM-T Easyvector(Promega ®)and subclonedin expression vector pCIneo, pCIL2B1generatingbuilding. The plasmid was transfectedinto 293 cellsin vitrofunctional analysisforthe expression ofL2. ObtainingconstructionpCIL2B1was confirmed by PCRand sequencing.The capacityof this construction to express the geneand produce their L2 protein inmammalian cellswas confirmed by RT-PCR and western blot (usinganti body against the epitopeAU1). The results confirmed the L2 gene expression in mammalian cellsand the consequent production of the protein L2 BPV-1.

Orientadores: Marcelo Menossi Teixeira, Rodrigo Franco de Carvalho / Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-25T18:01:39Z (GMT). No. of bitstreams: 1 Modolo_DiegoGrando_D.pdf: 3351414 bytes, checksum: 2f6c8310f0efbb6e6f3d2e48e34de879 (MD5) Previous issue date: 2014 / Resumo: O Brasil é conhecido por ter o segundo maior efetivo bovino e por ser o maior exportador de carne bovina no mundo. Dentre os problemas que afetam a pecuária nacional está a papilomatose bovina, uma doença infecto contagiosa causada pelo papilomavírus bovino (BPV). Diversas doenças de alto impacto econômico não só na pecuária nacional mas também mundial estão associadas ao BPV, sendo que ainda não existe uma vacina que possa prevenir o alastramento da doença e nem métodos eficázes de tramento. A proteína L1 do capsideo do BPV tipo 1 é uma forte candidata para ser utilizada na formulação vacinal por ser altamente imunogênica e ter a capacidade de formar, sozinha, partículas "virus-like" (VLPs). Devido a incapacidade de se multiplicar papilomavírus "in vitro", a utilização de sistemas de produção de proteínas recombinantes é a melhor estratégia para produção de proteínas virais em larga escala que possam ser utilizadas em formulações vacinais ou em testes de diagnósticos. Neste trabalho desenvolvemos uma plataforma de produção e purificação da proteína recombinante L1 em bactéria. A proteína L1 recombinante foi purificada por cromatografia de troca iônica e foi possível obter frações em alto nível de pureza. Também produzimos plantas transgênicas visando a produção da proteína L1. A transformação genética de plantas foi confirmada por PCR e RT-PCR, mas devido a inespecificidade dos anticorpos comerciais disponíveis e a uma possível baixa expressão do gene L1 em plantas, não foi possível confirmar a expressão da proteína recombinante. A proteína L1 obtida em bactérias poderá ser analisada como vacina e bem como na obtenção de anticorpos mais específicos e na produção de testes de diagnósticos. O conhecimento obtido neste trabalho poderá ser adaptado no desenvolvimento de outras vacinas de importância socio-econômica. / Abstract: Brazil is known as the largest exporter of beef and for having the second largest cattle herd in the world. Several illness that affect the national cattle industry are associated to the bovine papillomatosis, an infectious disease caused by Bovine Papillomavirus (BPV). Althought the economic impacts of these diseases affect the livestock at a global scale, there is no BPV vaccine or effective treatment methods available yet. The L1 capsid protein of BPV type 1 is a good candidate to be used in a vaccine formulation due its high immunogenicity and the ability to form virus-like particles (VLPs). Due to the inability to multiply papillomavirus in vitro, the use of recombinant protein production systems is the best strategy to produce viral proteins in large scale that can be used in vaccine formulations or for diagnosis testing. In this work, we developed a bacteria expression system to produce the recombinant L1 protein. The recombinant L1 protein was purified by ion exchange chromatography and highly purified fractions of L1 were obtained. We also obtained transgenic plants to produce the L1 protein. The genetic transformation of plants was confirmed by PCR and RT-PCR. Due to lack of specificity of commercial antibodies used in this study and a possible low expression of the L1 gene in plants, it was not possible to confirm the accumulation of the recombinant protein. The protein obtained in bacteria can be evaluated as vaccine as well as in the production of more specific antibodies and diagnosis tests. The knowledge obtained in this work can be adapted in the development of vaccines for other important socio-economic diseases / Doutorado / Genetica Vegetal e Melhoramento / Doutor em Genetica e Biologia Molecular

The bovine papillomavirus type 1 E2 protein is a multifunctional early viral protein with roles in all phases of the cell cycle. E2 is required during G1 as a transcription factor, in S phase to initiate viral replication and during mitosis to tether the viral genome to dividing DNA. The viral genome contains 17 E2 binding sites, the majority of which are concentrated in the long control region (LCR), a regulatory region that is upstream of the viral coding sequence. The role of these binding sites has been explored in vitro using small plasmids and E1 and E2 proteins expressed in bacteria and insect cells. In this study we attempt to examine the placement of E2 on its binding sites during all phases of the cell cycle and in the context of a stably replicating viral system. As part of the examination of the role of E2 during mitosis, we have also examined the role of the cohesin protein Scc1 in viral tethering. Two groups have published disparate reports identifying the cellular protein that binds to the transactivation domain of E2 to stably maintain viral genomes during cell division. Our group has published that it is the DNA helicase ChlR1 that is required for viral tethering, while it has been reported that it is the bromodomain protein Brd4 that is responsible. In this study we contribute to a report that shows that the cellular protein Scc1 binds to the viral genome through a ChlR1 independent mechanism. The cohesin protein binds to BPV-1 E2 at intermittent stages of the cell cycle and may be a factor in viral genome tethering. This interaction may also be important for regulating viral transcription.

Submitted by Erika Demachki (erikademachki@gmail.com) on 2017-05-05T19:00:03Z No. of bitstreams: 2 Dissertação - Winnie Castro Amorim e Albuquerque - 2017.pdf: 2279498 bytes, checksum: 160e4a276e405c6393dd5bb7742b106b (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2017-05-10T13:37:01Z (GMT) No. of bitstreams: 2 Dissertação - Winnie Castro Amorim e Albuquerque - 2017.pdf: 2279498 bytes, checksum: 160e4a276e405c6393dd5bb7742b106b (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2017-05-10T13:37:01Z (GMT). No. of bitstreams: 2 Dissertação - Winnie Castro Amorim e Albuquerque - 2017.pdf: 2279498 bytes, checksum: 160e4a276e405c6393dd5bb7742b106b (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2017-03-31 / Outro / Bovine papillomavirus is the etiological agent of bovine papillomatosis, a disease that triggers warts throughout the skin, udder, roofs, genitalia and in more severe cases can develop extensive papillomas, cause neoplasia in the digestive tract and bladder, weaken the animal's health and cause losses in the Productivity and losses for livestock. The present study aims to detect and typify bovine Papillomavirus present in bovine tissue and blood samples with papillomatosis, to sequence the isolated viral types, to analyze the nucleotide sequences and the phylogeny of the detected viral types. As a result, amplification was obtained in five tissue samples (papilloma) from different bovines, not being successful in the amplification of blood samples. PCR reactions revealed the presence of BPV-1 in 60%, BPV-5 in 40%, BPV-9, BPV-10, BPV-13 and BPV-14 in 20% and BPV-12 in 40% of the analyzed samples. The presence of coinfection was verified in 60% of the lesions analyzed, with up to four viral types infecting the same sample. Alignments of viral type sequences 1, 5 and 14 were validated with identity ranging from 74% to 95%. The phylogenetic diagram showed a genetic approximation between viral types 1 and 14, both belonging to the genus Deltapapillomavirus, and distancing between nucleotide sequences of viral types 5, 9 and 14. Papillomaviruses of types 5 and 9 belong to different genera, Epsilonpapillomavirus and Xipapillomavirus, Respectively, the phylogenetic distance between these viral types, verified in the diagram, is justified. / Papilomavírus bovino é o agente etiológico da papilomatose bovina, doença que desencadeia verrugas por toda pele, úbere, tetos, genitália e em casos mais graves pode desenvolver papilomas extensivos, causar neoplasia no trato digestivo e bexiga, debilitar a saúde do animal e provocar perdas na produtividade e prejuízos para a pecuária. O presente estudo possui como objetivo detectar e tipificar Papilomavírus bovino presentes em amostras de tecido e sangue de bovinos com papilomatose, sequenciar os tipos virais isolados, analisar as sequências nucleotídicas e a filogenia dos tipos virais detectados. Como resultado, obteve-se amplificação em cinco amostras de tecido (papiloma) de diferentes bovinos, não obtendo sucesso na amplificação das amostras de sangue. As reações de PCR revelam a presença do BPV-1 em 60%, BPV-5 em 40%, BPV-9, BPV-10, BPV-13 e BPV-14 em 20% e BPV-12 em 40% das amostras analisadas. A presença de coinfecção foi verificada em 60% das lesões analisadas, com até quatro tipos virais infectando a mesma amostra. Os alinhamentos das sequencias do tipo viral 1, 5 e 14 foram validados com identidade variando de 74% a 95%. O diagrama filogenético demonstrou aproximação genética entre os tipos virais 1 e 14, ambos pertencentes ao gênero Deltapapillomavirus, e distanciamento entre as sequências nucleotídicas dos tipos virais 5, 9 e 14. Os papilomavírus dos tipos 5 e 9 pertencem a gêneros diferentes, Epsilonpapillomavirus e Xipapillomavirus, respectivamente, justifica-se dessa forma, a distância filogenética entre esses tipos virais, verificada no diagrama.

The aim of this study was to develop a system for easy production of different veterinary chimeric vaccines based on stable mouse polyomavirus (MPyV) structures. The system is designed for antigens that are problematic in production or stability. First, universal vectors for baculovirus-directed production of chimeric MPyV VLPs or pentamers based on the major capsid protein VP1 were designed to be exploited as vaccines against other pathogens. The different strategies used in this study are based on: A) exposure of selected immunogenic epitopes on the surface of MPyV VLPs by inserting them into a surface loop of the VP1 protein, B) insertion of foreign protein molecules inside the VLPs, or C) fusion of a foreign protein or its part with the C-terminus of VP1 protein, thus forming giant pentamers of a chimeric protein. Candidate vaccine antigens against porcine circovirus 2 (PCV2), the causative agent of porcine circovirus 2 systemic diseases (PCV2-SD) which causes significant economic losses in swine breeding, were prepared using the constructed vectors. All candidate vaccines induced the production of antibodies against the capsid protein of PCV2 after immunization of mice. The candidate vaccine Var C based on fusion of MPyV and PCV2 capsid proteins, is able to induce production of antibodies with...

Thirty bovine with neoplasms of the upper digestive tract (UDT) associated to the spontaneous consumption of bracken fern (Pteridium aquilinum) were studied. They were from 27 farms, localized in the municipalities of Jaguari (23) and Nova Esperança do Sul (4), Rio Grande do Sul, Brazil. The total bovine population in those farms was 1,090 and large amounts of bracken fern were found in the pastures. Twenty-six were cows e four were castrated males. The age ranged from 3 to 13-years-old. Most of them were 7 to 8-years-old (46,6%). Clinical signs observed in the affected animals were progressive weight loss, absence of ruminal movements, cough, dysphagia, regurgitation, halitosis, diarrhea, and bloat. Less frequent signs were selective appetite, dyspnea, and salivation. Two bovines died and 28 were euthanatized in extremis and submitted to necropsy. The main gross and microscopic findings were found in the same areas of the UDT. They were digestive papillomatosis, transforming papillomas, and squamous cell carcinomas (SCC). Twenty-nine bovines had papillomas of various sizes in several areas of the UDT. The digestive papillomatosis ranged from mild (45%), to moderate (38%), to severe (17%). Three developing phases were observed microscopically in the examined papillomas: an early growing phase, a developing phase, and a regressing phase. The regressing phase was characterized by lymphocytic infiltrates at the base of the papilloma. In 16 cases, the microscopic examination of lesions grossly resembling papillomas (although some were slightly round, with lower or ulcerated finger-like projections) revealed malignant transformation of the papillomas into SCCs. The SCCs were solitary (12/30) or multiple (18/30) and were histologicaly characterized as well, moderately, or poorly differentiated. Grouping the distribution of SCCs of larger extension in the UDT into cranial region (base of the tongue, pharynx/oropharynx, and epiglottis), medial region (esophagus), and caudal region (cardia and rumen), the distribution was cranial in 39%, middle in 16%, and caudal in 45% of the cases. By the same grouping criteria, but considering the total number of times SCCs of varied extensions were diagnosed in the cranial, middle, and caudal regions, the percentages changed to 34%, 26%, and 40%, respectively. Metastases to regional lymph nodes and other organs, like liver and lungs, were observed in 18 cases. Immunohistochemistry for cytokeratin was performed in selected sections of SCCs and metastases, showing strong positive reaction in the well and moderately differentiated SCCs, but weak positive reaction in the poorly differentiated ones. The epidemiological and histomorphological evidences showed in this study are in agreement with the observations that point out the co-carcinogesis between bovine papillomavirus type 4 infection and chemicals of bracken fern in the pathogenesis of the SCCs in the UDT of cattle. / Foram estudados 30 bovinos com neoplasias no trato alimentar superior (TAS) associadas ao consumo espontâneo de samambaia (Pteridium aquilinum) provenientes de 27 propriedades rurais, sendo 23 no município de Jaguari e quatro em Nova Esperança de Sul, Rio Grande do Sul. A população bovina total das 27 propriedades em que ocorreram os casos era de 1.090 bovinos e havia quantidade abundante de samambaia nas áreas de pastoreio dos animais. Vinte e seis bovinos eram vacas e quatro eram machos castrados. A idade variou de três a 13 anos, sendo o maior número de casos entre sete e oito anos (46,6%). Os sinais clínicos observados incluíram emagrecimento progressivo, atonia ruminal, tosse, disfagia, regurgitação, halitose, diarréia e timpanismo. Outros sinais clínicos menos freqüentes foram apetite seletivo, dispnéia e salivação. Dois bovinos tiveram morte espontânea e 28 foram submetidos à eutanásia in extremis e necropsiados. Os principais achados macroscópicos e histológicos observados nos 30 bovinos localizavam-se nos mesmos locais do TAS e consistiam de papilomas, papilomas em transformação para carcinomas de células escamosas (CCEs) e CCEs. Vinte e nove bovinos tinham papilomas de diversos tamanhos, sendo a quantidade variável entre leve (45%), moderada (38%) e acentuada (17%). Nos papilomas examinados microscopicamente, foram observadas três fases de desenvolvimento: a) fase inicial de crescimento; b) fase de desenvolvimento; e c) fase de regressão; essa última era caracterizada por infiltrados linfocitários nos eixos fibrovasculares de sustentação. Em 16 bovinos, o exame histológico de lesões macroscópicas compatíveis com papilomas, porém alguns deles apresentando-se mais arredondados, com projeções digitiformes atenuadas ou ulceradas, revelou a transformação maligna desses papilomas em CCEs. Os CCEs eram únicos (12/30) ou múltiplos (18/30) e variaram quanto ao grau de diferenciação celular entre bem diferenciados, moderadamente diferenciados ou pouco diferenciados. Quando a distribuição dos CCEs de maior extensão foi agrupada em regiões cranial (base da língua, faringe/orofaringe, epiglote), média (terços cranial, médio e caudal do esôfago) e caudal (entrada do rúmen e rúmen) do TAS, observou-se que a localização era cranial em 39% dos casos, média em 16%, e caudal em 45%. Utilizando-se esse mesmo critério de agrupamento, porém considerando o número total de vezes em que CCEs (de extensões variadas) foram diagnosticados nas regiões cranial, média e caudal, os números alteraram-se para 34%, 26% e 40%, respectivamente. Metástases de CCEs para linfonodos regionais e outros órgãos como fígado e pulmão foram observadas em 18/30 bovinos. A técnica de imunoistoquímica para citoqueratina foi realizada em cortes selecionados de CCEs e metástases, observando-se células fortemente positivas nos CCEs bem e moderadamente diferenciados, e fraca imunomarcação nos pouco diferenciados. As evidências epidemiológicas e histomorfológicas relatadas neste estudo reforçam as observações de uma estreita correlação entre a infecção pelo papilomavírus bovino tipo 4, causador da papilomatose digestiva, e a co-carcinogênese química dos princípios tóxicos da samambaia na patogênese dos CCEs do TAS de bovinos.

Human papillomavirus infection is the cause of more than 99% of cervical cancer cases. The current vaccine is ineffective therapeutically; highlighting the need for continued papillomavirus research. One avenue that could be explored in this regard is the function of the papillomavirus E2 regulatory proteins. HPV E2 represses expression of the viral E6 and E7 oncoproteins. Reintroduction of E2 into cervical carcinoma cells results in growth arrest and cellular senescence. Understanding the mechanism of how E2 regulates the early promoter may be key to developing new therapeutic and prophylactic vaccines. Here, we describe regulation of E2 through acetylation and possibly through direct interaction with a novel cellular interacting protein, RINT1. Histone acetyltransferase (HAT) proteins have been demonstrated to interact with Bovine Papillomavirus (BPV) and Human Papillomavirus (HPV) E2 proteins as well as enhance E2 dependant transcription luciferase reporter plasmid containing E2 binding sites. We demonstrate that HATs p300, CBP, and pCAF are limiting for E2 dependant transcriptional activation and that each protein functions independently. We have also identified that BPV-1 E2 is a substrate for acetylation by p300. Mutants of E2 that cannot be acetylated on lysines 111 or 112, display abnormal transcriptional phenotypes. Cells deficient in p300 display similar transcriptional defects that are intensified by CBP depletion. We propose that acetylation of BPV-1 E2 is necessary for transcriptional activation. Acetylation generates a binding site through which a co-factor may interact via a bromodomain. Regulation of E2 dependent transcriptional activation through a post-transcriptional modification represents a novel method through which BPV-1 controls gene expression. We also present evidence for a direct interaction between BPV-1 E2 and the cellular factor RINT1. This interaction does not appear to be critical for transcriptional regulation; however, several other functional pathways are indicated by the cellular complexes in which RINT1 functions. Some of these, such as ER/Golgi vesicular transport and hTERT independent telomere maintenance, are pathways in which E2 has no known role. Further investigation into regulation and consequences of E2 acetylation and the biological significance of the interaction between E2 and RINT1 could prove important in understanding the complex role of E2 in papillomavirus infection.