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SUMMARY:Regulation of thiamine biosynthesis in Chlamydomonas reinhardtii -
  Zetty Northana Balia Yusof
DTSTART:20100507T120000Z
DTEND:20100507T123000Z
UID:TALK23960@talks.cam.ac.uk
CONTACT:15560
DESCRIPTION:\nABSTRACT In bacteria\, many genes involved in the biosynthes
 is of cofactors such as TPP are regulated by riboswitches\, a part of an m
 RNA molecule that can directly bind a small target molecule. Upon binding\
 , these RNA elements undergo conformational changes which then affect the 
 gene's activity. In the green alga Chlamydomonas reinhardtii\, we have fou
 nd that TPP riboswitches are found in THI4 and THIC\, the genes encoding t
 he first enzymes of the thiazole and pyrimidine branches respectively of t
 he thiamine biosynthesis pathway. The transcripts of these genes are found
  to be alternatively spliced when thiamine is added to the cultures and it
  is suggested that the presence of the riboswitches may play a role in thi
 s (Croft et al.\, 2007). The focus of my research has been to study how th
 iamine biosynthesis is regulated in C. reinhardtii mutants\, especially th
 e pyr1 mutant. The pyr1 mutant of C. reinhardtii (Smyth et al.\, 1975)\, w
 hich is resistant to the thiamine analogue pyrithiamine\, has a mutation i
 n the THI4 riboswitch that prevents the THI4 gene from being repressed by 
 TPP. I have analysed the growth and expression of thiamine biosynthesis ge
 nes in wild type and C. reinhardtii pyr1 mutants in the presence of thiami
 ne and pyrithiamine. Using a construct in which the THI4 riboswitch is clo
 ned upstream of the luciferase gene\, I have carried out in vivo expressio
 n analysis of the effect of mutations in conserved bases located in P2 and
  P4 stems of the C. reinhardtii THI4 riboswitch. I observed high expressio
 n of luciferase activity without the presence of thiamine\, and low expres
 sion with the presence of thiamine in the non-mutated riboswitch. When a s
 ingle base located in the P2 stem was changed to mimic the pyr1 mutant\, t
 he luciferase activity was maintained even in the presence of thiamine. A 
 single base change of one the conserved residues in the P4 stem also resul
 ted in a stable luciferase activity in the presence of thiamine. The resul
 ts confirm the participation of bases located in P2 and P4 stems in the re
 cognition of TPP as well as the consequence of binding of TPP towards the 
 expression of THI4 gene in C. reinhardtii. 
LOCATION:Department of Plant Sciences\, Large Lecture Theatre
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