EXPRESSION AND FUNCTIONAL ANALYSIS OF THE CALMODULIN GENE IN BLASTOCLADIELLA EMERSONII

Systematic functional analysis of calcium‐signalling proteins in the genome of the rice‐blast fungus,Magnaporthe oryzae, using a high‐throughput RNA‐silencing system

“…Interestingly, sporulation and appressorium formation were severely affected in the latter mutant despite the low level of silencing, indicating that CAM plays a crucial role in those biological processes. By pharmacological approaches using CAM antagonists such as W7, W5, R24571, TFP, chlorpromazine, 48/80 and calmidazolium, involvement of CAM in growth and/or infection structure developments were also shown previously in various fungi (Roy and Datta, 1987; Magalhaes et al ., 1991; Robson et al ., 1991; Warwar and Dickman, 1996; Rao et al ., 1997; Lee and Lee, 1998; Liu and Kolattukudy, 1999; Simao and Gomes, 2001; Singh et al ., 2001; Judelson and Roberts, 2002; Uhm et al ., 2002; Ahn et al ., 2003; Carvalho et al ., 2003; Chung, 2003; Sato et al ., 2004; Ahn and Suh, 2007). Our results provided genetic evidence that supports the findings of previous pharmacological studies.…”

Blastocladiella emersonii expresses a centrin similar to Chlamydomonas reinhardtii isoform not found in late‐diverging fungi

“…The migration pattern observed in SDS-PAGE with the recombinant BeCen1 protein in the presence of calcium did not show the large mobility shift expected for four Ca 2+ -binding domains, like that obtained for calmodulin [22]. According to the consensus sequence of the Ca 2+ -binding sites [31], only the third EF-hand domain was not expected to bind calcium because of a threonine at position 5 and an asparagine at position 12.…”

“…Total RNA was extracted from synchronized cells at different stages of B. emersonii developmental cycle, using the Trizol method (Life Technologies), and Northern blots were performed as previously described [22] with some modifications. RNA was transferred to Hybond N+ membranes (Amersham) and EcoRI/NotI fragments from the sequenced library clones were used as probes.…”

Expressão gênica diferencial durante a esporulação de Blastocladiella emersonii e estudo da sinalização por GMP cíclico

“…Até bem recentemente, o conhecimento sobre os genes expressos em B. emersonii limitava-se aos genes dos RNAs ribossômicos e a oito seqüências codificadoras de proteínas (de Oliveira et al, 1994;de Souza and Gomes, 1998;Fietto et al, 2002;Rocha and Gomes, 1999;Simao and Gomes, 2001;Stefani and Gomes, 1995;Van der Auwera and De Wachter, 1996).…”

“…Simão & Gomes (2001) isolaram e caracterizaram o gene que codifica a proteína calmodulina em B. emersonii. Análises por Northern blot mostraram que os níveis do mRNA de calmodulina são regulados ao longo do desenvolvimento de B. emersonii, permanecendo baixo nas células vegetativas e aumentando durante a esporulação (Simao and Gomes, 2001). Com a obtenção de anticorpos policlonais foi possível mostrar que os níveis da proteína calmodulina (CaM) seguem o mesmo padrão do seu mRNA (Simao and Gomes, 2001).…”