Intellectual merit: Biotin is an essential molecule, whose primary function has been ascribed as a catalytic cofactor required by enzymes involved in diverse metabolic processes. Recent studies indicate that biotin is also a regulatory molecule that appears to play critical roles in controlling transcriptional and post-transcriptional mechanisms in gene expression. This research will identify and characterize the biochemical and physiological functions of genes associated with the biotin metabolic network of eukaryotic organisms, using Arabidopsis as a model. Plants are ideally suited for these studies as they, along with some microbes, are the primary organisms that can synthesize this molecule de novo; all other organisms must acquire this molecule from their diets or from the environment. This project will characterize aspects of the biotin metabolic network that are unique to the organism under study, namely Arabidopsis, as a model plant. Specifically: 1) how do developmental and environmental cues regulate biotin biosynthesis; these studies will take advantage of the two biotin biosynthetic genes that have recently been identified by the investigators, BIO1 and BIO2; and 2) what is the biochemical and physiological function of plant-specific biotin-containing proteins. These are methylcrotonyl-CoA carboxylase (encoded by genes At1g03090 and At4g34030), the seed-specific biotin protein (encoded by gene At2g42560), and three biotin carrier-like proteins (encoded by genes At1g52670, At3g15690, At3g56130); and 3) what are the mechanisms by which biotin regulates gene expression at both the transcriptional and post-transcriptional level. Broader impact: The project will be a vehicle for the education and training of a new cadre of scientists at the undergraduate, graduate and post-graduate levels. In-lab research experiences will be provided for exposing young scientists to multidisciplinary basic research in functional genomics and biotechnology. The research will define the organization of a complex metabolic network that is structured around the catalytic and regulatory functions of biotin. This achievement will provide novel insights into how such complex networks are regulated. The goal is to develop an example of how to view metabolism as a complex integrated network rather than the classical textbook model of a set of linear pathways. The development of this view of metabolism is possible only in the context of an organism whose genome is completely sequenced. Such an understanding of metabolism is required in order to comprehensively understand how metabolic processes are interactively regulated by developmental and environmental cues.
