The Binder of SPerm 1 (BSP1) protein is involved in the fertilization and semen cryopreservation processes and is described to be both beneficial and detrimental to sperm. Previously, the relationship of BSP1 with freezability events has not been completely understood. The objective of this work was to determine the differential abundance of the forms of the BSP1 protein in cryopreserved seminal plasma of Bos taurus indicus bulls with different patterns of semen freezability using proteomics. A wide cohort of adult bulls with high genetic value from an artificial insemination center was used as donors of high quality, fresh semen. Nine bulls presenting different patterns of semen freezability were selected. Two-dimensional gel electrophoresis showed differential abundance in a group of seven protein spots in the frozen/thawed seminal plasma from the bulls, ranging from 15 to 17 kDa, with pI values from 4.6 to 5.8. Four of these spots were confirmed to be BSP1 using mass spectrometry, proteomics, biochemical, and computational analysis (Tukey's test at P < 0.05). The protein spot weighing 15.52 ± 0.53 kDa with a pI value of 5.78 ± 0.12 is highlighted by its high abundance in bulls with low semen freezability and its absence in bulls presenting high semen freezability. This is the first report showing that more than two forms of BSP1 are found in the seminal plasma of Nelore adult bulls and not all animals have a similar abundance of each BSP1 form. Different BSP1 forms may be involved in different events of fertilization and the cryopreservation process.

Asian soybean rust, caused by the fungus Phakopsora pachyrhizi, is one of the most important diseases affecting soybean production in tropical areas. During infection, P. pachyrhizi secretes proteins from haustoria that are transferred into plant cells to promote virulence. To date, only one candidate P. pachyrhizi effector protein has been characterized in detail to understand the mechanism by which it suppresses plant defenses to enhance infection. Here, we aimed to extend understanding of the pathogenic mechanisms of P. pachyrhizi based on the discovery of host proteins that interact with the effector candidate Phapa-7431740. We demonstrated that Phapa-7431740 suppresses pathogen-associated molecular pattern triggered immunity (PTI), and it interacts with a soybean glucan endo-1,3- β-glucosidase (GmβGLU), a pathogenesis-related (PR) protein belonging to the PR-2 family. Structural and phylogenetic characterization of PR-2 protein family predicted in the soybean genome and comparison to PR-2 family members in Arabidopsis thaliana and cotton, demonstrated that GmβGLU is a type IV β-1,3-glucanase. Transcriptional profiling during an infection time course showed that the GmβGLU mRNA is highly induced during the initial hours after infection, coinciding with peak of expression of Phapa-7431740. The effector was able to interfere with the activity of GmβGLU in vitro, with a dose-dependent inhibition. Our results suggest that Phapa-7431740 may suppress PTI by interfering with glucan endo-1,3-β-glucosidase activity.

No abstract

Neosporosis has become a concern since it is associated with abortion in cattle. Currently, in situ diagnosis is determined through anamnesis, evaluation of the history, and perception of the clinical signs of the herd. There is no practical and noninvasive test adapted to a large number of samples, which represents a gap for the use of new approaches that provide information about infections and the risks of herds. Here, we performed a search in the Neospora caninum genome by linear B-cell epitopes using immunoinformatic tools aiming to develop a chimeric protein with high potential to bind specifically to antibodies from infected cattle samples. An enzyme-linked immunosorbent assay with the new chimeric antigen was developed and tested with sera from natural field N. caninum-infected bovines. The cross-reactivity of the new antigen was also evaluated using sera from bovines infected by other abortive pathogens, including Trypanosoma vivax, Leptospira sp., Mycobacterium bovis, and Brucella abortus, and enzootic bovine leucosis caused by bovine leukemia virus, as well as with samples of animals infected with Toxoplasma gondii. The assay using the chimeric protein showed 96.6% ± 3.4% of sensitivity in comparison to healthy animal sera. Meanwhile, in relation to false-positive results provided by cross-reactivity with others pathogens, the specificity value was 97.0% ± 2.9%. In conclusion, immunoinformatic tools provide an efficient platform to build an accurate protein to diagnose bovine neosporosis based on serum samples.

Enhanced photosynthesis is strictly associated with to productivity and it can be accomplished by genetic approaches through identification of genetic variation. By using a Solanum pennellii introgression lines (ILs) population, it was previously verified that, under normal (CO 2), IL 2-5 and 2-6 display increased photosynthetic rates by up to 20% in comparison with their parental background (M82). However, the physiological mechanisms involved in the enhanced CO 2 assimilation exhibited by these lines remained unknown, precluding their use for further biotechnological applications. Thereby, here we attempted to uncover the physiological factors involved in the upregulation of photosynthesis in ILs 2-5 and 2-6 under normal (CO 2) as well as under elevated (CO 2). The results provide evidence for increased biochemical capacity (higher maximum carboxylation velocity and maximum electron transport rate) in plants from IL 2-5 and 2-6, whereas the diffusive components (stomatal and mesophyll conductances) were unaltered in these ILs in comparison to M82. Our analyses revealed that the higher photosynthetic rate observed in these ILs was associated with higher levels of starch as well as total protein levels, specially increased RuBisCO content. Further analyses performed in plants under high (CO 2) confirmed that biochemical properties are involved in genetic variation on chromosome 2 related to enhanced photosynthesis.

A produção de glicoproteínas terapêuticas possui alto custo devido a necessidade de utilização de cultura de células de mamíferos que dependem de meios de cultivo complexos e caros e possuem baixo rendimento de produção. Leishmania tarentolae, um protozoário tripanossomátideo não patogênico para mamíferos, tem sido sugerido como um sistema alternativo para expressão heteróloga de glicoproteínas devido a existência de métodos eficientes de expressão heteróloga, ser facilmente adaptado para produção em alta escala e depender de meios de cultura de baixo custo. Além disso, este protozoário apresenta modificações pós- traducionais ausentes em bactérias e leveduras, organismos mais utilizados para produção industrial de proteínas recombinantes. O Interferon-β, é uma proteína utilizada em tratamentos de doenças como esclerose múltipla, as suas glicosilações são extremamente relevantes para suas funções biológicas. Embora o perfil de glicosilação de proteínas expressas em L. tarentolae seja semelhante a mamíferos, existem algumas diferenças no perfil de carboidratos presentes nas terminações das cadeias glicídicas devido à ausência de enzimas de biossínteses e incorporação de ácido siálico. Nesta tese foi desenvolvido e otimizado uma plataforma de expressão de glicoproteínas utilizando linhagens de L. tarentolae geneticamente otimizadas para síntese de glicoproteínas em alta escala mais estáveis com incorporação de ácido siálico na extremidade da cadeia glicídica e sua aplicação na produção de Interferon-β humano. Além da validação da produção do Interferon-β, também foi realizado caracterização de seus glicanos e validação de sua atividade para regulação de inflamação em modelo murinho, demonstrando que esta nova linhagem geneticamente otimizada de L. tarentolae é adequada para produção em larga escala de glicoproteínas terapêuticas. Palavras-chave: Leishmania tarentolae. Interferon-β. Proteínas terapêuticas. Glicosilação

Horses are seasonal polyoestrous animals, and the photoperiod is the main factor modulating their reproductive activity. There is no consensus on the andrological and biochemical factors that influence breeding seasonality. To assess the involvement of climate in reproduction, Mangalarga Marchador stallions were monitored over 1 year regarding semen quality and seminal plasma proteome. Here, we show that kallikrein (KLKs) proteoforms in seminal plasma are involved in climate conditioning of reproduction. During the breeding season, greater abundance and different types of KLKs occurred simultaneously to lower sperm motility, greater semen volumes and higher concentrations of glucose and cholesterol. Considering that vasodilation due to activation of the kallikrein-kinin system and the consequent inhibition of the renin-angiotensin system may be associated with lower sperm motility, unravelling the involvement of KLK proteoforms in reproductive seasonality is a priority in horse breeding.