Gene expression in cotton (Gossypium hirsutum L.) fiber: cloning of the mRNAs.

Abstract

Cotton, an important natural fiber, is a differentiated epidermal cell. The number of genes that are active in fiber cells is similar to those in leaf, ovule, or root tissues. Through differential screening of a fiber cDNA library, we isolated five cDNA clones that are preferentially expressed in fiber. One of the cDNA clones, pCKE6, corresponded to an abundant mRNA in fiber. Transcripts for E6 were detected throughout the development of the fiber. Immunoprecipitation of in vitro translation products and Western blot analysis of fiber proteins showed two polypeptides in the range of 30-32 kDa as the products of E6 mRNA. Sequence analysis and hybrid-selected RNA translation also suggest that E6 mRNAs encode two polypeptides. Concentrations of E6 mRNA and protein are highest during the late primary cell wall and early secondary cell wall synthesis stages. Sequence comparison of E6 with other known eukaryotic and prokaryotic genes reveals no significant homology (GenBank; December 1991). E6 or a homologous gene(s) is conserved in several members of Malvaceae as well as in one other fiber-producing plant, kapok, but is not found in several other plants examined or in Acetobacter xylinum. A genomic clone corresponding to pCKE6 was isolated, and the promoter element of the E6 gene was shown to direct the expression of a carrot extensin mRNA in a tissuespecific and developmentally regulated fashion in transgenic cotton plants.Cotton is the most important textile fiber crop; therefore, the development of cotton fiber and the biosynthesis of cellulose are significant areas of research. Cotton fiber is a differentiated single epidermal cell of the outer integument of the ovule. It has four distinct growth phases; initiation, elongation (primary cell wall synthesis), secondary cell wall synthesis, and maturation (for a review, see ref. 1). Initiation of fiber development appears to be triggered by hormones, auxins, and gibberellins (for a review, see ref.2). The primary cell wall is laid down during the elongation phase, lasting up to 25 days postanthesis (DPA). Synthesis of the secondary wall commences prior to the cessation of the elongation phase and continues to -40 DPA, forming a wall (5-10 ,um) ofalmost pure cellulose. Upon maturity, cotton fiber is -87% cellulose. Thus cell wall biosynthesis is a major synthetic activity in fiber cells. The cell wall components are synthesized and transported by a functionally integrated membrane system of endoplasmic reticulum, Golgi complex, and plasmalemma. Newly synthesized cell wall polypeptides are released into the endoplasmic reticulum lumen before transportation and incorporation into cell walls. Glycosylation of structural proteins, as well as polymerization of hemicellulose and pectin, takes place in the Golgi complex from which the products are released into plasmalemma through the fusion of vesicles (1, 3).Plants, as well as certain bacteria, synthesize cellulose through enzyme systems localized in plasma membranes that use UDP-glucose as a substrate (1). The en...

The Cotton Transcription Factor TCP14 Functions in Auxin-Mediated Epidermal Cell Differentiation and Elongation

“…The cDNAs of cotton genes were amplified by RT-PCR from total RNAs of cotton root, hypocotyl, leaf, flower, ovules, and fiber cells according to the ultracentrifugation method (John and Crow, 1992). The cDNAs of Arabidopsis genes were amplified by RT-PCR from total RNA extracted from the seedlings using TRIzol reagent (Invitrogen).…”

Isolation of genes preferentially expressed in cotton fibers by cDNA filter arrays and RT-PCR

“…Sequencing of 5?-terminal ends identified 30 different genes. A BLASTX search of the NCBI database identified several fiber-genes already reported, including those encoding E6 [2], expansin [8], annexin [23] and several cytoskeletal components, such as actin, a-and b-tubulins. However, there were a number of newly discovered cDNAs (Table 2).…”

“…Since the first report of isolation of several cDNAs that showed a higher expression in developing fiber cells [2], a few dozen fiber-genes were reported and their expression patterns analyzed [3,4]. A number of these genes, such as H6 , Rac13, Fb12A , FS5/6 and GhEX1 , are considered fiber-specific [5 Á/8], whereas others, such as E6 , B6 , and LTP6 , are fiber-enriched [2,9 Á/11].…”

Trichome Specific Expression: Promoters and Their Applications

“…Initially, fiber specific genes were isolated by differential screening of cDNA library. This led to the successful identification of several genes with strong and specific expression in fibers, including E6, genes encoding Lipid Transfer Proteins (LTPs), a Proline Rich Protein and other genes with no obvious sequence similarity (John and Crow, 1992;Ma et al, 1995;Orford and Timmis, 1995;Rinehart et al, 1996;Orford and Timmis, 1997;Orford and Timmis, 1998;Orford et al, 1999). Already, Northern or RT-PCR analysis showed that genes can be expressed during distinct phases of development of the fiber cells or throughout the life of these cells.…”