טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
Ph.D Thesis
Ph.D StudentHacham Yael
SubjectRegulation of Methionine Metabolism in Plants by the
N-Terminal Region of Cystathionine gamma-synthase
DepartmentDepartment of Biology
Supervisors Professor Gadi Schuster
Professor Rachel Amir
Full Thesis textFull thesis text - English Version


Abstract

Methionine biosynthesis is regulated by cystathionine gamma-synthase (CGS), the first specific enzyme of its pathway. Researchers have found that the level of CGS is controlled by the MTO1 domain located within the N-terminal region, and that this regulation is mediated by the level of S-adenosylmethionine (SAM), a methionine metabolite. Our major objective was to identify additional domains within the N-terminal region that are important for CGS regulation. We found another form of CGS mRNA with 90 nt deletion within the N-terminal region. This deleted form of CGS was found in vivo in Arabidopsis, in addition to the full-length CGS. Its association with polyribosomes indicates that this form is translated in cells. Unlike the full-length CGS, the deleted form of CGS is not feedback-regulated by methionine

Methionine, lysine and threonine belong to the aspartate amino acid family. Our second objective was to further study the crosstalk between methionine and threonine, and to explore the effect of enhanced carbon/amino skeleton flux towards methionine synthesis on methionine level in plant cells. To this end, we crossed transgenic tobacco plants overexpressing feedback-insensitive bacterial aspartate kinase (AK) that contains a significantly higher threonine level with transgenic plants overexpressing different forms of AtCGS (full-length, 90 nt deleted form or the truncated CGS without the N- terminal region). We found that while threonine didn’t affect the expression level of the full-length CGS it significantly enhanced the expression level of the deleted form of CGS, suggesting that this form maintains a high level of methionine when a high level of threonine is produced.

In order to study how a high level of lysine affects methionine synthesis, we crossed between plants overexpressing the feedback-insensitive bacterial enzyme dihydrodipicolinate synthase (DHPS), which contains a significantly higher level of lysine, and plants overexpressing full-length AtCGS. Significantly higher methionine levels accumulated in plants expressing both genes compared to plants expressing CGS alone, while the lysine level remained similar to that overexpressing DHPS alone. We found a new regulatory pathway where an increased lysine level results in reduced levels of SAM synthase and therefore reduced levels of SAM. Since SAM is a negative regulator of CGS, this reduction leads to a higher CGS expression and consequently to an increased methionine level.

Elucidating the relationship between methionine and lysine or methionine and threonine may open up new ways of producing transgenic crop plants containing increased levels of these amino acids and thus improve their nutritional quality.