Abstract
Chinese cabbage (Brassica campestris) is an economically important leaf vegetable crop worldwide. Mounting studies have shown that cysteine-cysteine-cysteine-histidine (CCCH) zinc-finger protein genes are involved in various plant growth and development processes. However, research on the involvement of these genes in male reproductive development is still in its infancy. Here, we identified 11 male fertility-related CCCH genes in Chinese cabbage. Among them, a pair of paralogs encoding novel non-tandem CCCH zinc-finger proteins, Brassica campestris Male Fertility 30a (BcMF30a) and BcMF30c, were further characterized. They were highly expressed in pollen during microgametogenesis and continued to express in germinated pollen. Further analyses demonstrated that both BcMF30a and BcMF30c may play a dual role as transcription factors and RNA-binding proteins in plant cells. Functional analysis showed that partial bcmf30a bcmf30c pollen grains were aborted due to the degradation of pollen inclusion at the microgametogenesis phase, and the germination rate of viable pollen was also greatly reduced, indicating that BcMF30a and BcMF30c are required for both pollen development and pollen germination. This research provided insights into the function of CCCH proteins in regulating male reproductive development and laid a theoretical basis for hybrid breeding of Chinese cabbage.
Highlights
The eukaryotic genome encodes a large number of proteins containing a variety of zinc-finger motifs and forms one of the most abundant protein families in eukaryotes
The results suggested that the mutated Brassica campestris Male Fertility 30a (BcMF30a) and BcMF30c may still encode the peptide chains, but their synthesis may be terminated prematurely, and it is predicted that the newly synthesized peptide chains lack the C-terminal of the full-length proteins containing the RNA recognition motif (RRM) domain (Figure 6A)
Subcellular localization analysis showed that they can be localized to the nucleus, the signals of both BcMF30a/BcMF30c-eGFP and eGFP-BcMF30a/BcMF30c fusion proteins in the nucleus were weaker than those of free-eGFP (Figure 4 and Figure S3). These findings demonstrated that BcMF30a and BcMF30c may function as nuclear transcriptional activators in plant cells
Summary
The eukaryotic genome encodes a large number of proteins containing a variety of zinc-finger motifs and forms one of the most abundant protein families in eukaryotes. In this study, we screened several male fertility-related CCCH zinc-finger protein genes by identifying the differentially expressed genes between the ‘Bcajh97-01A/B’ genetic male sterile (GMS) A/B lines of B.campestris Among them, both Brassica campestris Male Fertility 30a (BcMF30a) and its paralog BcMF30c encode novel non-TZF proteins. We identified 11 CCCH zinc-finger protein genes that may be involved in male fertility in B. campestris They are all expressed during pollen development, they display different expression patterns and differ in gene and protein structures. The results suggested that the mutated BcMF30a and BcMF30c may still encode the peptide chains, but their synthesis may be terminated prematurely, and it is predicted that the newly synthesized peptide chains lack the C-terminal of the full-length proteins containing the RRM domain (Figure 6A).
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