The availability of the
carbon backbone
O-phosphohomoserine (OPHS) is critical to
methionine (met) and
threonine (thr) synthesis. OPHS derives from
homoserine and is formed by
homoserine kinase (HSK). To clarify the function of HSK in cellular metabolism, the E. coli HSK ortholog thrB was expressed in potato plants targeting the EcHSK
protein to chloroplasts and to the cytosol. Both approaches resulted in up to 11 times increased total HSK
enzyme activity. Transgenic plants exhibited reduced
homoserine levels while met and thr did not accumulate significantly. However, the precursor
cysteine and upstream intermediates of met such as
cystathionine and
homocysteine did indicating an accelerated
carbon flow towards the end products. Coincidently, plants with elevated cytosolic levels of EcHSK exhibited a reduction in transcript levels of the endogenous HSK, as well as of
threonine synthase (TS),
cystathionine beta-lyase (CbL), and met synthase (MS). In all plants,
cystathionine gamma-synthase (CgS) expression remained relatively unchanged from wild type levels, while
S-adenosylmethionine synthetase (
SAMS) expression increased. Feeding studies with externally supplied
homoserine fostered the synthesis of met and thr but the regulation of synthesis of both
amino acids retained the wild type regulation pattern. The results indicate that excess of plastidial localised HSK activity does not influence the de novo synthesis of met and thr. However, expression of HSK in the cytosol resulted in the down-regulation of gene expression of pathway genes probably mediated via OPHS. We integrated these data in a novel working model describing the regulatory mechanism of met and thr homeostasis.