Protein N-terminal acetylation (Nt-acetylation) is the transfer of acetyl group from acetyl coenzyme A (Ac-CoA) to the alpha amino acid of a protein. Since it has been discovered more than fifty years ago, Nt-acetylation is known to be one of the most common protein modifications in eukaryotes, occurring on approximately 50-70% of yeast soluble protein and about 80-90% of human protein. However, the exact biological role has remained enigmatic for majority of affected proteins, and only for a small number of proteins, Ntacetyation was linked to various features of protein such as localization, stability and interaction. Nt-acetylation in yeast and in human is thoroughly investigated with the identification of five (NatA-NatE) and six (NatA-NatF) Nα-acetyltransferase (NAT) types, respectively. In contrast, the knowledge of Nt-acetylation in plants was vacant for many years. The first Arabidopsis NAT, AtNatC was identified in 2003, and very recently three more NATs (NatA, NatB and NatE) were described by Iwona Stephan. In this study, we identified two NATs (NatD and NatF) that are still missing in plants. AtNatD/AtNaa40p is conserved from yeast with respect to acetylation of protein histone H4. The lack of Nterminal serine acetylation increases the overall positive charge of H4 N-tail which causes the minor phenotypes observed in atnaa40 mutant. The acetylation of N-terminal serine of histone H4 might also involve in DNA double-strand break response. Besides, the subcellular localization to cytoplasm and nucleus suggests a lysine acetyltransferase activity of AtNaa40p towards histones. AtNatF/AtNaa60 unusually localizes to plasma membrane and to the tonoplast. The sensitivity of atnaa60 mutant to salt tress during germination stage appears to be related to the localization, and indicating the involvement of AtNaa60p in salt stress or osmotic stress response. Like hNaa60p, AtNaa60 is believed to acetylate a large number of proteins according to the NBD-Cl fluorescent assay. AtNaa60p acetylates methione and serine starting peptides in vitro. In addition, numerous proteins are found N-terminally acetylated in chloroplasts, both chloroplast-encoded and nuclear-encoded proteins. In silico study reveals eight putative plastidic NATs of which seven localize to the chloroplasts when they are transiently expressed with EYFP in Arabidopsis protoplasts. Three proteins (At2g39000, At1g24040 and At2g06025) acetylate plenty of Escherichia coli proteins, their substrate specificities are strongly correlated to chlotoplast transit peptide (cTP) cleavage sites. Four other proteins (At4g19984, At1g26220, At1g32070 and At4g28030) are possibly true NATs since they possess the conserved Ac-CoA binding motif. Our results, together with other studies on acetylation in chloroplast, propose the connection between Nt-acetylation of chloroplastic proteins and drought stress.
|Supervisor:||Hell, Prof. Dr. Rüdiger|
|Date of thesis defense:||16 December 2013|
|Date Deposited:||13 Jan 2014 07:59|
|Faculties / Institutes:||Service facilities > Centre for Organismal Studies Heidelberg (COS)|
|Subjects:||570 Life sciences|