General regulatory factor 3 regulates the expression of alternative oxidase 1a and the biosynthesis of glucosinolates in cytoplasmic male sterile Brassica juncea.

Mitochondrial retrograde signaling (MRS) plays an essential role in sensing and responding to internal and external stimuli to optimize growth to adapt to the prevailing environmental conditions. Previously studies showed alterations on MRS in cytoplasmic male sterile (CMS) plant. However, the regulators involved in MRS in CMS plants remain largely unknown. In this study, we used alternative oxidase 1a (AOX1a) as an indicator of MRS and found that the expression of AOX1a was significantly downregulated in a CMS line comparing to its revertant line, thus indicating an alteration in MRS in the CMS line. By performing a BLAST search of known regulatory components involved in MRS in yeast, we identified general regulatory factor 3 (GRF3), an orthologue of Bmh1/2 in yeast, and demonstrated an association between this gene and MRS in plants, as evidenced by change in AOX1a expression. GRF3 protein was found to be located in the nucleus and the plasma membrane. Further studies showed that GRF3 interacted with MYB29, and regulated the biosynthesis of glucosinolates in Brassica juncea. These findings revealed that GRF3, a negative regulator of AOX1a, is involved in MRS, and also plays a vital role in the accumulation of glucosinolates in CMS crops.

The casein kinase 2 ß subunit CK2B1 is required for swollen stem formation via cell cycle control in vegetable Brassica juncea.

The swollen stem is a determinant of yield for the stem-type vegetable Brassica juncea that is representative of vegetative organ formation. However, the genetic mechanism underlying swollen stem formation and its regulation remains unknown. In this study, we identified a casein kinase 2 ß subunit 1 (CK2B1) and revealed its role in swollen stem formation. Genotyping analysis revealed that a homozygous variation in the CK2B1 promoter is responsible for swollen stem formation, and the promoter activity of CK2B1 was significantly associated with the variations between swollen stem and non-swollen stem types. CK2B1 was exclusively located in the nucleus and expressed in the stem nodes of the plant. Swollen stem formation was blocked when CK2B1 expression was silenced, and induced in a backcross population carrying a swollen stem genotype, which indicates that CK2B1 is required for swollen stem formation. Cell numbers were increased during swollen stem formation and decreased in CK2B1-silenced expression plant, indicating that CK2B1 regulates swollen stem formation via cell division. CK2B1 directly interacted with E2Fa, a regulator of G1/S transition in the cell cycle, in which CK2 phosphorylates E2Fa. Our results revealed that CK2B1 affects swollen stem formation via the control of the cell cycle. These findings help to elucidate the signals that control swollen stem formation and provide a promising molecular target to enhance the yield of vegetative organ formation.