Abstract
Maize, a salt-sensitive crop, frequently suffers severe yield losses due to soil salinization. Enhancing salt tolerance in maize is crucial for maintaining yield stability. To address this, we developed an introgression line (IL76) through introgressive hybridization between maize wild relatives Zea perennis, Tripsacum dactyloides, and inbred Zheng58, utilizing the tri-species hybrid MTP as a genetic bridge. Previously, genetic variation analysis identified a polymorphic marker on Zm00001eb244520 (designated as ZmSC), which encodes a vesicle-sorting protein described as a salt-tolerant protein in the NCBI database. To characterize the identified polymorphic marker, we employed gene cloning and homologous cloning techniques. Gene cloning analysis revealed a non-synonymous mutation at the 1847th base of ZmSC(IL76) , where a guanine-to-cytosine substitution resulted in the mutation of serine to threonine at the 119th amino acid sequence (using ZmSC(Z58) as the reference sequence). Moreover, homologous cloning demonstrated that the variation site derived from Z. perennis. Functional analyses showed that transgenic Arabidopsis lines overexpressing ZmSC(Z58) exhibited significant reductions in leaf number, root length, and pod number, alongside suppression of the expression of genes in the SOS and CDPK pathways associated with Ca(2+) signaling. Similarly, fission yeast strains expressing ZmSC(Z58) displayed inhibited growth. In contrast, the ZmSC(IL76) allele from Z. perennis alleviated these negative effects in both Arabidopsis and yeast, with the lines overexpressing ZmSC(IL76) exhibiting significantly higher abscisic acid (ABA) content compared to those overexpressing ZmSC(Z58) . Our findings suggest that ZmSC negatively regulates salt tolerance in maize by suppressing downstream gene expression associated with Ca2+ signaling in the CDPK and SOS pathways. The ZmSC(IL76) allele from Z. perennis, however, can mitigate this negative regulatory effect. These results provide valuable insights and genetic resources for future maize salt tolerance breeding programs.