Increased endogenous brassinosteroid level promotes fiber cell elongation by over-expressing GhDWF4 gene in transgenic cotton (Gossypium hirsutum L.)

Functional Genomics

oral

Luo, Ming; Hu, Mingyu; Asami, Tadao; Nakano, Takeshi; Fujioka, Shozo ; Pei, Yan; Xiao, Yuehua

Luo, Ming; Hu, Mingyu; Asami, Tadao; Nakano, Takeshi; Fujioka, Shozo ; Xiao, Yuehua

Pei, Yan

Cotton is the leading fiber crop in the world, producing the most prevalent natural fibers used in textile industry. Cotton fibers are single cells derived from outer epidermis of ovules. Their growth and development consist of four overlapping stages: fiber initiation, cell elongation, secondary cell wall deposition, and maturation. Fiber cells from G. hirsutum are 1000 to 3000 times longer than they are wide. With highly elongated structure composed almost of cellulose, cotton fibers are considered as an ideal model for studies of plant cell elongation and cell wall biogenesis. Brassinosteroids (BRs) are a kind of steroidal hormone in plant kingdom. Exogenous application of BRs increased fiber length in field experiments or in ovule culture condition. Contrarily, brassinazole (BRZ), a specific inhibitor of BR biosynthesis, repressed cotton fiber initiation and elongation. However, the role of endogenous BRs and the mechanism of BRs activity are largely unknown in the growth and development of cotton fiber cell. To illuminate the effect of endogenous BRs on fiber cell development, we have cloned GhDWF4 gene, a DWF4 (steroid C-22 hydroxylase) homologue from upland cotton and generated a lot of transgenic cotton lines over-expressing GhDWF4 or suppressing GhDWF4 expression. In the transgenic fiber cell over-expressed GhDWF4 gene, the endogenous BRs content increased. In 12-DPA transgenic fiber cell, for example, 6-Deoxocathasterone, 6-deoxotyphasterol, and 6-Deoxocastasterone increased by 53 fold, 3 fold, and 4 fold, respectively. In 20-DPA transgenic fiber cell, 6-Deoxocathasterone, 6-deoxoteasterone, 6-Deoxotyphasterol, and 6-Deoxocastasterone increased by 46 times, 22 times, 25 times, and 7 times, respectively. Meanwhile, the changes have not detected for the intermediates involved in the early C-6 oxidation pathway between transgenic fiber and control. The fiber length of CaMV35S::GhDWF4 increased by 9.8%-17.14% and the fiber length of pFBP7::GhDWF4 increased by 4.5%-7.7% compared with control. Furthermore, the strength of GhDWF4-increased fibers improved while micronaire decreased. The fiber strength of pFBP7::GhDWF4 and CaMV35S::GhDWF4 increased 2.8% and 9.8%, respectively. These result revealed that enhanced endogenous BRs level not only promoted fiber cell elongation but also improved the quality of cotton fiber. To understand the molecular mechanism of BRs promoting fiber elongation, we performed digital gene expression analysis used 12-DPA fibers. The results indicated that large numbers of important genes related to fiber elongation were up-regulated, such as aquaporins and cellulose biosynthases. It is suggested that increased BRs level promotes many physiological processes involved in fiber cell elongation. In addition to fiber length increased, the growth and development of transgenic plants were modified. The plants overexpressing GhDWF4 grew faster and displayed larger leaf size, longer stalks, and bigger flowers and balls. GhDWF4 repressed plants displayed dwarf, deep green and small leaves, and shorter internodes. The growth of a few lines was seriously suppressed, displaying severely dwarf and very slow growth. The height of 2-year grown plant was still shorter than 20cm, and the leaf stalk length and internode length were extremely short. GhDWF4–repressed plants displayed sterile phenotype for flower bud abortion.