Functional characterization of chlorophyll b reductase NON-YELLOW COLORING 1 in Medicago truncatula

Chlorophyll degradation is a characteristic process of leaf senescence. Two mutant lines, which showed green leaves and seeds during senescence, were identified by screening a Tnt-1 retrotransposon-tagged population of Medicago truncatula. Genetic and molecular analyses indicated that the mutated gene is NON-YELLOW COLORING 1 (MtNYC1) in M. truncatula. MtNYC1 encoded a chlorophyll b reductase, characterized by three transmembrane domains and a catalytic site (Y***K). Our investigation further identified three splicing variants of MtNYC1, encoding a full-length protein (MtNYC1A) and two truncated proteins (MtNYC1B, MtNYC1C). Genetic evidence indicated that the catalytic site and the third transmembrane domain were critical domains for chlorophyll b reductase. The coordinated action of three splicing variants plays a pivotal role in the degradation of chlorophyll during the senescence of leaves. This discovery provides precise target sites for the development of stay-green legume cultivars.