Cloning and Expression Analysis of Two Cotton (Gossypium Hirsutum L.) Genes Encoding Cell Wall Proline-rich Proteins

Abstract

Two cotton (Gossypium hirsutum L.) genes, ghprpl and ghprpl, encoding cell wall proline-rich proteins (PRPs) have been cloned and characterized. The ghprpl gene has an open reading frame (ORF) that encodes a PRP of 299 amino acids (aa), whereas the ghprp2 gene contains an ORF that codes for a 310-aa PRP. The GhPRPl has an 80% identity in aa sequence with that of GhPRP2. Like other plant cell wall PRPs, both cotton PRPs have a hydrophobic signal peptide at their N-termini, followed by repeating peptide units. Northern blot analyses showed that the ghprp1 gene is predominantly expressed in the fiber during the elongation stage of fiber development. Reverse transcription (RT)-PCR analysis showed that ghprp1 is expressed in both fiber and root tissues, whereas ghprpl is in roots only. The ghprp1 gene was shown to be present in the A1, A2, D1 and D5 genomes of Gossypium by PCR amplification, whereas the ghprpl gene is only present in the A1 and A2 genomes. The ghprp1 gene was over-expressed in the yeast Pichia pastoris, and the expressed GhPRP1 protein was used as an antigen to raise polyclonal antibodies (anti-GhPRP1). Western analysis using the anti-GhPRP1 probe detected a major protein band (50 kDa) in 5-31-day postanthesis (DPA) fibers. However, the 50 kDa protein was absent in other cotton tissues.