Poster: Reproductive development

Abs # 375: A Lily Pollen-Specific and Desiccation-Associated cDNA Encoding the CRIB Motif-Containing Protein

Presenter:	Hsu, Ssu-Wei , ssuwei0112@pchome.com.tw
Authors	Hsu, Ssu-Wei  (A)   Wang, Co-Shine  (A)
Affiliations:	(A): Graduate Institute of Biotechnology, National chung Hsing University

This work characterizes a pollen-specific and desiccation-associated transcript that encodes the CRIB motif-containing protein in lily (Lilium longiflorum Thunb. cv. Snow Queen) pollen during development and stress. The transcript, designated LLP12-2, encodes a gene product having a sequence of 222 amino acids, a calculated molecular mass of 24 kDa, and a calculated pI of 9.1. Assessment of the hydropathy shows that the polypeptide is hydrophilic but it is not a heat-stable protein. The protein contains a high content of glycine, serine and proline (11-16%) with relatively high amounts (8%) of arginine and lysine. Sequence analysis demonstrates similarities between the predicted lily LLA12-2 and a family of conserved Cdc42/Rac-interactive binding (CRIB) motif-containing proteins that may interact with Cdc42/Rac GTPases. Antiserum was raised against the overexpressed LLP12-2 protein in Escherichia coli. Affinity-purified LLP12-2 antibodies were prepared from antiserum to investigate the specificity and distribution of the protein during development. The affinity-purified LLP12-2 antibodies recognized both the 32- and 35-kDa pollen proteins and the two polypeptides were immunologically related. Immunoblot analyses of total protein from floral and vegetative organs confirmed that LLP12-2 proteins accumulated to detectable levels only in a discrete stage of anther development. Premature drying of developing pollen indicated that the accumulation of LLP12-2 proteins was associated with desiccation. Subcellular fractionation of pollen proteins revealed that the protein was located in the cytosolic fraction.