Cotton Fiber Preferentially Expressed In The Preliminary Study Of The Abc Transporter Protein

Cotton fibers, which are single cells derived from seed coat, provide a good system for investigating cell fate determination, cell elongation and cellulose synthesis. A cDNA library was constructed which contains genetic messages necessary for early cotton fiber development. 105 ESTs were isolated and sequenced. Expression of the 15 ESTs in intact ovules of Xu142 and glabrous ovules of fl mutant was analyzed. It was found that one EST was preferentially expressed in the intact ovules of Xu142. Its corresponding gene, GhABC1, encodes an ATP-binding cassette (ABC) transporter protein. Low levels of transcripts were also detected in other organs as well as mutant ovules by RT-PCR and Northern hybridization. The expression of GhABC1 was up-regulated in fibers during the fiber elongation stage. In li mutant ovules expression levels of GhABC1, E6 and Expansin genes were low, even in the early stage of cotton fiber development. Southern blot indicated that there are at least three copies of GhABC1 gene in the diploid genome. Three transgenic vectors were constructed to transform cotton, one for overexpression, the second for antisense RNA and the third for double-strand RNA to repress the endogenous gene transcription. The fiber-specific promoter E6 was used for all the three vectors. To determine the subcellular localization of GhABC1 protein, GFP fused with GhABC1 was generated. The construct was transformed into onion epidermal cells. Transient expression indicated that GhABC1 was localized in the plasm membrane.The overexpression (OE) and dominant-negative mutant (DN) forms driven by a constitutive 35S promoter have been introduced into Arabidopsis thaliana. The transformants showed interesting phenotypic changes. Both OE and DN lines have some plants with very short siliques (SS). RT-PCR analysis reveals that AtWBC11, a Arabidopsis homolog of GhABC1, was predominantly expressed in reproductive organs, such as siliques and inflorescence. The expression level of endogenous AtWBC11 gene in the transgenic and wild type Arabidopsis plants is comparable. However, higher levels of GhABC1 transcripts were detected in the siliques of SS lines than in the siliques of normal OE transgenic lines. A T-DNA insertion line of AtWBC11 has been obtained from Arabidopsis Biological Resource Center (ABRC). Homozygous T-DNA lines of AtWBC11 exhibit fused leaves and short siliques. Although have flowers, they did't produce seeds. Characterization of AtWBC11 will help to dissect the function of GhABC1 protein.Cotton (Gossypium hirsutum L.) li mutant has shortened fibers (seed-hairs), curling leaves and distorted organ appearance. Observation with SEM did not reveal clear difference in fiber initiation and early growth between the wild-type and li mutant ovules. However, during elongation stage (e.g., 3 days post anthesis, DPA), the fiber of li mutant showed little growth, and is significantly shorter than the fiber of wild-type. It is well known that the distribution pattern of axuin plays important roles in regulating plant development including organ bilateral symmetry formation and vascular tissues differentiation. An assay of polarauxin transport showed that, in comparison with wild type plants, the basipetal transport activity was greatly reduced in the stem of li mutant. Stem transection showed an incomplete differentiation and development of cambium and phloem in li mutant, a phenomenon consistent with its reduction in polar auxin transport. These results suggest that phenotypic changes in li mutant, including early stop of fiber elongation, are likely related to impaired polar auxin transport.