Uma família multigênica, polimórfica, ainda pouco conhecida de P. falciparum é a família SURFIN, codificada por dez genes denominados surf. Para abordar o modo de transcrição desses genes e observar se ocorre switching transcricional, monitoramos a quantidade de transcritos analisando amostras por RT-qPCR ao longo de 40 reinvasões. O gene surf 4 foi claramente detectado em nossos ensaios, apresentando quantidade significativa de transcritos na fase esquizonte. Para analisar a localização e função de SURFIN 4, construímos linhagens transgênicas (por fusão à GFP) e por fusão de um domínio desestabilizador (DD24) que permite de forma controlada, desestabilizar a proteína e analisar a sua importânica. Não houve alteração na viabilidade dos parasitas com SURFIN 4 desestabilizado, no entanto detectamos um aumento significativo de transcritos quando SURFIN 4 foi diminuída. Estes achados apontam para um novo mecanismo de regulação gênica, ao menos para o gene surf 4 onde a proteína SURFIN 4 parece ser capaz de modular a quantidade do seu respectivo transcrito. / One smaller family of antigen variant of P. falciparum whose function is still unknown is surf genes. Some members appear in asexual blood stage forms and may be associated also to virulence associated processes. We accessed the transcription of the members of the surf gene family along multiple invasions by real time PCR. Based on the observation of constitutive expression of one surf gene, surf4.1, we created a parasite line which expresses a GFP, conditionally destabilized SURFIN4.1 protein. Upon destabilization of the protein, no differential growth rates were observed. However, when we monitored the transcription of SURFIN4.1 knocked-down parasites compared to their stabilized counterparts, we observed a strong increase in the transcript quantities of surf4.1 and also three other surf genes, pointing to a feedback of the SURFIN protein quantity to either the stability of surf transcripts or the transcriptional activity of surf loci. This type of potential regulation is yet unheard in Plasmodium biology.

Uma família multigênica, polimórfica, ainda pouco conhecida de P. falciparum é a família SURFIN, codificada por dez genes denominados surf. Para abordar o modo de transcrição desses genes e observar se ocorre switching transcricional, monitoramos a quantidade de transcritos analisando amostras por RT-qPCR ao longo de 40 reinvasões. O gene surf 4 foi claramente detectado em nossos ensaios, apresentando quantidade significativa de transcritos na fase esquizonte. Para analisar a localização e função de SURFIN 4, construímos linhagens transgênicas (por fusão à GFP) e por fusão de um domínio desestabilizador (DD24) que permite de forma controlada, desestabilizar a proteína e analisar a sua importânica. Não houve alteração na viabilidade dos parasitas com SURFIN 4 desestabilizado, no entanto detectamos um aumento significativo de transcritos quando SURFIN 4 foi diminuída. Estes achados apontam para um novo mecanismo de regulação gênica, ao menos para o gene surf 4 onde a proteína SURFIN 4 parece ser capaz de modular a quantidade do seu respectivo transcrito. / One smaller family of antigen variant of P. falciparum whose function is still unknown is surf genes. Some members appear in asexual blood stage forms and may be associated also to virulence associated processes. We accessed the transcription of the members of the surf gene family along multiple invasions by real time PCR. Based on the observation of constitutive expression of one surf gene, surf4.1, we created a parasite line which expresses a GFP, conditionally destabilized SURFIN4.1 protein. Upon destabilization of the protein, no differential growth rates were observed. However, when we monitored the transcription of SURFIN4.1 knocked-down parasites compared to their stabilized counterparts, we observed a strong increase in the transcript quantities of surf4.1 and also three other surf genes, pointing to a feedback of the SURFIN protein quantity to either the stability of surf transcripts or the transcriptional activity of surf loci. This type of potential regulation is yet unheard in Plasmodium biology.

The interactions between Medicago truncatula and Phytophthora medicaginis were examined using skl, a mutant blocked in ethylene perception, and a range of wild accessions of this plant species. P. medicaginis infection of M. truncatula plants resulted in compatible responses, whereas the mutant genotype was found to be hyper-susceptible to the pathogen. Phytophthora reproduction and colonization rates of Medicago tissues supported this conclusion. Infection of skl with different pathogens reinforced this observation. Ethylene production in infected A17 and skl roots showed reduced ethylene evolution in the mutant and suggested that a positive feedback loop, known as autocatalytic ethylene production, amplified the ethylene signal. To complement the study, expression analyses of defense response genes in this interaction were studied by real time RTPCR of Phytophthora-infected and mock-infected roots. The genes analyzed were PAL, CHS, IFR, ACC oxidase, GST, and PR10. The sequences needed for the analysis were found through the scrutiny of the M. truncatula EST database employing phylogenetics and bio-informatics tools. In A17 all the genes studied were up-regulated, although the specific gene expression patterns differed. The comparison of gene expression between A17 and skl genotypes allowed the differentiation between ethylene-dependent and ethylene-independent responses. Discrete results showed that ACC oxidase homologues were downregulated in the ethylene perception mutant, corroborating the ethylene observations. However, the expression of genes involved in the phenylpropanoid metabolism was increased in skl relative to A17, suggestive of an antagonism between the ethylene perception pathway and the regulation of the phenylpropanoid pathway. This result implied that Medicago phytoalexins accumulate in the disease interaction, but raised questions about their role in resistance to Phytophthora infection. This study establishes a link between mechanisms that regulate symbiotic infection and the regulation of disease resistance to Oomycete pathogens, especially P. medicaginis. The results served to identify a series of Phytophthora-induced genes, which remain pathogen-responsive even in the absence of a functional ethylene perception pathway. While it is possible that the products of these genes are involved in resistance to P. medicaginis, the present results demonstrate that ethylene perception is required for resistance.