Genomic and functional characterization of polyamine pathway genes in soybean.

摘要

Polyamines are currently targeted in chemotherapeutic intervention in cancer research. The role of soybean as a dietary chemoprevention factor increases because of its potential to reduce endogenous polyamine content and inhibit tumorogenesis. We investigated genomic organization and expression of polyamine associated genes and polyamine content to create resources for future investigation of polyamine function and possible modification of polyamines in soybean.; The putrescine pathway and spermidine/spermine pathway, including methionine biosynthesis, were investigated. Expressed sequenced tags (ESTs) survey provided an effective starting point to gain information on organization and expression of polyamine associated genes. Among approximately 310,000 ESTs in EST libraries, diverse representation of polyamine associated genes were identified, eg: seven for cystathionine beta lyase (CBL) and spermine synthase (SPMS) and 618 for S-adenosylmethionine synthetase ( SAMS). EST frequencies of polyamine associated genes corresponded to steady-state transcript levels determined using reverse transcriptase polymerase chain reaction (RT-PCR). Southern analysis demonstrated that polyamine associated genes are present in small to medium gene families in the soybean genome. Using bacterial artificial chromosome libraries, genome coverage up to 40X, physical loci were identified for putrescine biosynthesis genes: arginine decarboxylase (ADC) and ornithine decarboxylase ( ODC) and for spermine biosynthesis genes: S-adenosylmethionine decarboxylase (SAMDC), spermidine synthase (SPDS), and SPMS. Close physical linkage, within 150 kb, was detected for ADC, putrescine biosynthesis, and diamine oxidase (DAO), putrescine catabolism, in both of the duplicated loci in soybean genome.; Spermidine was most abundant in seeds and leaves, whereas putrescine was highest in juvenile roots. Spermine was the least abundant polyamine. Real Time RT-PCR detected higher steady-state levels of ADC than ODC in roots, leaves, shoot apices, and seeds, suggesting ADC is the predominant pathway for putrescine biosynthesis in soybean.; The soybean fasciation mutant, characterized by enlarged apical meristem and accelerated primordia formation in early postembryonic development, had three fold higher putrescine content in seeds compared with several wild-type soybean genotypes. High putrescine content and hastened spermine biosynthesis during phase II germination may be important for establishment of mutant phenotype. Following heavy metal exposure, down regulation of DAO indicated that putrescine, not spermidine and spermine, is critical for cellular protection in soybean.