Abstract
The 14-3-3 gene family members play key roles in various cellular processes. However, little is known about the numbers and roles of 14-3-3 genes in wheat. The aims of this study were to identify TaGF14 numbers in wheat by searching its whole genome through blast, to study the phylogenetic relationships with other plant species and to discuss the functions of TaGF14s. The results showed that common wheat harbored 20 TaGF14 genes, located on wheat chromosome groups 2, 3, 4, and 7. Out of them, eighteen TaGF14s are non-ε proteins, and two wheat TaGF14 genes, TaGF14i and TaGF14f, are ε proteins. Phylogenetic analysis indicated that these genes were divided into six clusters: cluster 1 (TaGF14d, TaGF14g, TaGF14j, TaGF14h, TaGF14c, and TaGF14n); cluster 2 (TaGF14k); cluster 3 (TaGF14b, TaGF14l, TaGF14m, and TaGF14s); cluster 4 (TaGF14a, TaGF14e, and TaGF14r); cluster 5 (TaGF14i and TaGF14f); and cluster 6 (TaGF14o, TaGF14p, TaGF14q, and TaGF14t). Tissue-specific gene expressions suggested that all TaGF14s were likely constitutively expressed, except two genes, i.e., TaGF14p and TaGF14f. And the highest amount of TaGF14 transcripts were observed in developing grains at 20 days post anthesis (DPA), especially for TaGF14j and TaGF14l. After drought stress, five genes, i.e., TaGF14c, TaGF14d, TaGF14g, TaGF14h, and TaGF14j, were up-regulated expression under drought stress for both 1 and 6 h, suggesting these genes played vital role in combating against drought stress. However, all the TaGF14s were down-regulated expression under heat stress for both 1 and 6 h, indicating TaGF14s may be negatively associated with heat stress by reducing the expression to combat heat stress or through other pathways. These results suggested that cluster 1, e.g., TaGF14j, may participate in the whole wheat developing stages, e.g., grain-filling (starch biosynthesis) and may also participate in combating against drought stress. Subsequently, a homolog of TaGF14j, TaGF14-JM22, were cloned by RACE and used to validate its function. Immunoblotting results showed that TaGF14-JM22 protein, closely related to TaGF14d, TaGF14g, and TaGF14j, can interact with AGP-L, SSI, SSII, SBEIIa, and SBEIIb in developing grains, suggesting that TaGF14s located on group 4 may be involved in starch biosynthesis. Therefore, it is possible to develop starch-rich wheat cultivars by modifying TaGF14s.
Highlights
The ubiquitous 14-3-3 proteins, as one of the families of regulatory proteins, have been found in all eukaryotic organisms and tissues
To explore the chromosomal locations and numbers of TaGF14 in wheat, the complete coding sequence of TaGF14-JM22 was used as the query to search the wheat whole-genome sequences published by IWGSC6
To investigate the evolutionary relationship among TaGF14JM22 and other GF14 genes and proteins derived from Oryza sativa, B. distachyon, Zea mays, Hordeum vulgare, and Arabidopsis thaliana, phylogenetic trees were constructed using the maximum likelihood method with a Poisson model and with 1000 bootstrap replicates by MEGA 6.0 (Tamura et al, 2013) based on the amino acid sequences of 14-3-3 proteins with a cut-off value of 50% for the condensed tree (Figure 2)
Summary
The ubiquitous 14-3-3 proteins, as one of the families of regulatory proteins, have been found in all eukaryotic organisms and tissues. The family consists of dimeric α-helical pSer/Thr binding proteins that play key roles in various cellular processes, such as signal transduction, biotic and abiotic stress responses, and carbon and nitrogen metabolism, by mediating protein– protein interactions (Aitken et al, 1992; Fulgosi et al, 2002). 14-3-3 proteins, binding a range of transcription factors and signaling proteins, have roles in regulating carbon and nitrogen metabolism, plant development, and biotic and abiotic stress responses (Roberts, 2000, 2003; Fulgosi et al, 2002; Maraschin et al, 2003). 14-3-3 proteins were reported to be involved in starch biosynthesis in plants. Only four GF14s have been reported in common wheat (Ikeda et al, 2000; Yao et al, 2005; Wang et al, 2008). Due to the roles of GF14 reported previously, it is necessary to study the 14-3-3 genes and their functions in wheat
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