Characterization of ScMat1, a putative TFIIH subunit from sugarcane.
The general transcription factor TFIIH is a multiprotein complex with different enzymatic activities such as helicase, protein kinase and DNA repair. MAT1 (ménage à trois 1) is one of the TFIIH subunits that has kinase activity and it is the third subunit of the cyclin-dependent kinase (CDK)-activating kinase (CAK), CDK7- cyclin H. The main objective of this work was to characterize ScMAT1, a sugarcane gene encoding a MAT1 homolog. Northern blots and in situ hybridization results showed that ScMAT1 was expressed in sugarcane mature leaf, leaf roll and inflorescence, and it was not differentially expressed in any of the other tissues analyzed such us bud and roots. In addition, ScMAT1 was not differentially expressed during different stress conditions and treatment with hormones. In situ hybridization analyses also showed that ScMAT1 was expressed in different cell types during leaf development. In order to identify proteins that interact with ScMAT1, a yeast two hybrid assay with ScMAT1 as bait was used to screen a sugarcane leaf cDNA library. The screening of yeast two hybrids yielded 14 positive clones. One of them is a cytochrome p450 family protein involved in oxidative degradation of toxic compounds. Other clones isolated are also related to plant responses to stress. To determine the subcellular localization of ScMAT1, a ScMAT1-GFP fusion was assayed in onion epidermal cell and the fluorescence was localized to the nucleus, in agreement with the putative role of ScMAT1 as a basal transcription factor.
Gentile
Agustina
A
Departamento de Genética e Evolução, Instituto de Biologia, Universidade Estadual de Campinas, Campinas, São Paulo, Brazil.
Ditt
Renata F
RF
Dias
Fabio O
FO
Da Silva
Marcio J
MJ
Dornelas
Marcelo C
MC
Menossi
Marcelo
M
D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D035264:Flowers / Q000235:genetics / Q000378:metabolism; D018506:Gene Expression Regulation, Plant; D015723:Gene Library; D017343:Genes, Plant; D008969:Molecular Sequence Data; D018515:Plant Leaves / Q000235:genetics / Q000378:metabolism; D010940:Plant Proteins / Q000235:genetics / Q000378:metabolism*; D018749:RNA, Plant / Q000378:metabolism; D031786:Saccharum / Q000235:genetics* / Q000378:metabolism; D016415:Sequence Alignment; D013312:Stress, Physiological; D051758:Transcription Factor TFIIH / Q000235:genetics / Q000378:metabolism*; D020798:Two-Hybrid System Techniques
D010940:Plant Proteins; D018749:RNA, Plant; D051758:Transcription Factor TFIIH