ReviewUntangling multi-gene families in plants by integrating proteomics into functional genomics
The classification and study of gene families is emerging as a constructive tool for fast tracking the elucidation of gene function. We review the growing role of proteomics in analysing gene families in model plant species by specifically identifying the products of closely related genes, determining their abundance, and coupled to affinity chromatography and subcellular fractionation studies, providing location within cells and functional assessment of specific family members.
Introduction
Historically, and for largely technical reasons, genes and their products have generally been studied as single entities. In this post-genomic era, with publicly available data from genome and EST sequencing projects, it is evident that most genes are not singletons but exist as members of gene families. Consequently, the polypeptide products of genes operate alongside similar proteins in cells that are encoded by other members of their gene families. The development of gene and protein specific technologies has paved the way for studying the expression and contribution of the individual members of such families and allows us to begin to answer questions about both partitioning of functions between members and redundancy within gene families. With the completions of both the Arabidopsis and rice genomes (Kaul et al., 2000; Goff et al., 2002) recent publications have utilised these genomic resources to build a picture of particular gene families. By taking a genomic perspective, researchers are able to define a gene family in the context of their gene of interest. The study of gene families will not only simplify the task of elucidating the function of every type of protein, but will also help us appreciate the evolutionary pressures leading to the expansion and preservation of gene duplicates within plant genomes. In this review we overview the scale of the gene family issue in model plants and consider research contributions that attempt to tackle it. We have not tried to systematically review all the literature, but give references to illustrate key aspects of gene family function and redundancy that have been discovered through experimentation to date. We especially consider the place of proteomic approaches, both applied and potential, in determining the expression, location and function of specific gene family products in plants.
Section snippets
Plant functional genomics – why study gene families?
John Donne placed individual people in the context of their society in the early 17th century when he wrote “no man is an island, entire of itself, every man is a piece of the continent, a part of the main” (Donne, 1999). The same concept can also be applied to our understanding of genes and proteins. Genes have their place and purpose in genomes and proteins their location and functional significance in the context of proteomes. The immediate influence of the wider genome and proteome on the
Making the most of gene specific technologies
The experimental study of gene families relies on the ability to readily distinguish between related members. Traditional methods for following gene expression, such as northern hybridisation, often failed to distinguish between the similarly sized members of gene families and suffered from cross-hybridisation between similar sequences. Only by separately following and identifying individual members of gene families and their products can we really address questions of differential expression,
Making the most of protein specific technologies
The annotation of genome sequences coupled with developments in mass spectrometry has meant that proteomics can now also be fully integrated into a genomic context and protein families can be explored. Rather than relying on a battery of specific antibodies to try and identify protein products, single mass spectrometers yield peptide mass fingerprints and tandem mass spectrometers deliver peptide sequence information to provide a high level of downstream specificity (Graves and Haystead, 2002).
Where to now? Challenges for the future in gene families
To date research of gene families has tended to focus on genomic organization of genes and some basic expression analysis. The next challenge is `functional' characterisation and it is probable that an intimate knowledge of the expression of gene families will greatly aid this process. This next stage will involve understanding the transcriptional regulation of related genes and appreciating the real and apparent levels of functional redundancy within defined genomes in plants. Detailed and
Acknowledgements
Research grants from the Australian Research Council Discovery Programme to A.H.M are greatly acknowledged. P.G.S is a recipient of a Grains Research and Development Corporation PhD scholarship and A.H.M is an Australian Research Council QEII Research Fellow.
Pia G. Sappl is a PhD student supported by a scholarship from the Australian Grain's Research and Development Corporation in the Plant Molecular Biology Group, School of Biomedical and Chemical Sciences at The University of Western Australia. She obtained her BSc (Hons 1st class) from The University of Western Australia, and was the recipient of the Lugg Medal for Biochemistry for her undergraduate degree. Her PhD research focuses on the glutathione S-transferase gene family in Arabidopsis,
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Pia G. Sappl is a PhD student supported by a scholarship from the Australian Grain's Research and Development Corporation in the Plant Molecular Biology Group, School of Biomedical and Chemical Sciences at The University of Western Australia. She obtained her BSc (Hons 1st class) from The University of Western Australia, and was the recipient of the Lugg Medal for Biochemistry for her undergraduate degree. Her PhD research focuses on the glutathione S-transferase gene family in Arabidopsis, using proteomic and genomic studies of the structure and expression of this family and T-DNA knock-out resources to probe gene family redundancy.
Joshua L. Heazlewood is Post-Doctoral Research Associate in the Plant Molecular Biology Group, School of Biomedical and Chemical Sciences at The University of Western Australia. He obtained his PhD in Plant Molecular Biology from La Trobe University, Australia investigating the MYB gene family in Arabidopsis using reverse genetic techniques. In 2001 he obtained a Post-Doctoral position with Dr. Millar's group at The University of Western Australia where he has been using liquid chromatography and gel electrophoresis coupled to tandem mass spectrometry to investigate the mitochondrial proteomes of model plants.
A. Harvey Millar is an Australian Research Council Queen Elizabeth II Research Fellow in the Plant Molecular Biology Group, School of Biomedical and Chemical Sciences at the University of Western Australia. He obtained his PhD in Biochemistry from the Australian National University, Canberra, Australia. He then worked as a Human Frontier Science Programme Long-Term Fellow in the Department of Plant Sciences in Oxford, UK, before returning to Australia in 1999 via a series of research fellowships held at The University of Western Australia. Dr. Millar's group is focussed on proteomic analysis in the model plants Arabidopsis and rice, with a special emphasis on mitochondrial proteomes and plant stress/defence strategies. His work aims to integrate proteomic data into biochemical analysis of plants, and also into the increasing information available in model plants relating to gene families, gene expression patterns and genetic resources.