Abstract
The polygalacturonase (PG) gene family has been found to be enriched in pollen of several species; however, little is currently known about the function of the PG gene in pollen development. To investigate the exact role that the PG gene has played in pollen development and about this family in general, one putative PG gene, Brassica campestris Male Fertility 9 (BcMF9), was isolated from Chinese cabbage (Brassica campestris ssp. chinensis, syn. B. rapa ssp. chinensis) and characterized. RT-PCR, northern blotting and in situ hybridization were used to analyse the expression pattern of BcMF9, and antisense RNA technology was applied to study the function of this gene. BcMF9 is expressed in particular in the tapetum and microspore during the late stages of pollen development. Antisense RNA transgenic plants that displayed decreased expression of BcMF9 showed pollen morphological defects that resulted in reduced pollen germination efficiency. Transmission electron microscopy revealed that the homogeneous pectic exintine layer of pollen facing the exterior was over-developed and predominantly occupied the intine, reversing the normal proportional distribution of the internal endintine layer and the external exintine in transgenic pollen. Inhibition of BcMF9 also resulted in break-up of the previously formed tectum and baculae from the beginning of the binucleate stage, as a result of premature degradation of tapetum. Several lines of evidence, including patterns of BcMF9 expression and phenotypic defects, suggest a sporophytic role in exine patterning, and a gametophytic mode of action of BcMF9 in intine formation. BcMF9 might act as a co-ordinator in the late stages of tapetum degeneration, and subsequently in the regulation of wall material secretion and, in turn, exine formation. BcMF9 might also play a role in intine formation, possibly via regulation of the dynamic metabolism of pectin.
Highlights
Studies of the Arabidopsis pollen transcriptome and pollen gene expression in other organisms have concluded that the reduced transcriptome of pollen accumulated a restricted and unique set of genes, including the extensive representation of gene families and genes related to specific pathways (Becker et al, 2003; Lee and Lee, 2003; Honys and Twell, 2004; Pina et al, 2005)
The corresponding full-length cDNA of Brassica campestris Male Fertility 9 (BcMF9) was obtained by RACE-PCR and the DNA sequence was amplified with the gene-specific primers designed according to the fulllength cDNA sequence
A homology search using the Sequence Retrieval System against the Swiss-Prot/TrEMBL database revealed that BcMF9 showed 86 % identity and 94 % similarity to the putative pollen-expressed PGA3 from Arabidopsis on the amino-acid level
Summary
Studies of the Arabidopsis pollen transcriptome and pollen gene expression in other organisms have concluded that the reduced transcriptome of pollen accumulated a restricted and unique set of genes, including the extensive representation of gene families and genes related to specific pathways (Becker et al, 2003; Lee and Lee, 2003; Honys and Twell, 2004; Pina et al, 2005). Several members of the ‘classical’ arabinogalactan gene family were detected exclusively in pollen, and have been indicated to play roles in pollen tube nutrition, pollen tube guidance and other processes (Cheung and Wu, 1999; Pereira et al, 2006) Members of another large family, the polygalacturonase (PG) gene family, were found to be expressed in pollen and/ or anthers of several species, including maize, tobacco (Nicotiana tabacum), cotton, Brassica napus and Arabidopsis (Allen and Lonsdale, 1993; Robert et al, 1993; John and Petersen, 1994; Tebbutt et al, 1994; Hadfield et al, 1998; Honys and Twell, 2003). BcMF9 might play a role in intine formation, possibly via regulation of the dynamic metabolism of pectin
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