2011 Annual Report 1a.Objectives (from AD-416) 1: To map and characterize leaf mutants in maize. [NP 301, C4, PS 4A] 2: To positionally clone genes that regulate plant architecture in maize. [NP 301, C4, PS 4A] 3: To determine the function of genes through transgenic tests. [NP 301, C4, PS 4A] 1b.Approach (from AD-416) Maize is an important crop as well as a model organism for other cereals such as sorghum, barley, rice and wheat. The large number of genetic mutants, in combination with the ease of obtaining and mapping additional mutants, makes maize an excellent system for determining the function of genes. We propose to identify genes that regulate maize leaf development and shoot architecture. We will carry out mutagenesis with defined inbreds using the chemical mutagen EMS. Mutants will be characterized genetically and histologically. The genes will be mapped to position and cloned. The functions will be determined by expression analysis and further genetics. We already have two mutants at different stages of analysis. The dominant Liguleless narrow mutant has been mapped to position and beginning characterization is under way. The dominant Wavy auricle in blade1 (Wab1) mutant has been localized to a BAC contig. Once we clone Wab1, we will determine the function of the wild-type gene product. In order to follow expression of the genes we clone, we have developed a vector for gene fusions. We are presently testing this vector with the liguleless1 gene. REPLACING 5335-21000-028-00D (09/10). 3.Progress Report ARS scientists in Albany, CA identified a mutant in a cell wall screen in maize that has high glucan levels in the cell walls. A mapping population was made and the region narrowed to 4 megabases on the long arm of chromosome 6. A candidate gene was identified and a base pair change identified in a conserved critical region of the protein. The gene encodes a lichenase. The KNOTTED1 (KN1) transcription factor functions to promote indeterminate cell identities and prevents determinate fates. We carried out chromatin immunoprecipitation to identify direct targets. We identified ~5000 genes that are targeted by KN1. Of interest is the fact that most hormone pathways are affected by KN1, suggesting it plays a critical role in hormone homeostasis. Studies on leaf mutants have progressed with mapping and characterization of the candidate genes. Hoja loca, a dominant mutant that produces pin-shaped meristems sometimes and wide leaves other times, maps to chromosome 8. It affects auxin transport. The candidate gene for liguleless narrow is a serine-threonine kinase. A duplicate gene exists in the grasses, which is upregulated in the dominant mutant. ragged leaf2 is a recessive mutant that has altered cell fate in leaves but no gross morphological problems. It has been mapped to chromosome. The wavy auricle in blade mutant was mapped to two genes. One is a transcription factor similar to terosinte branched, which restricts growth. This gene is upregulated in Wab mutants, suggesting it is the correct gene. 4.Accomplishments 1. Identification of a maize mutant (candy leaf-1) with increased glucose in the cell wall. An impediment to plant feedstocks is obtaining sugars from the cell wall due to the fact they are embedded in a matrix of lignin and mixtures of C5 and C6 sugars, which are hard to ferment. ARS scientists at Albany, CA, in collaboration with scientists at UC Berkeley identified a maize mutant with increases in glucose in the cell wall in both seedling and adult leaves. The mutant was mapped to a lichenase gene, which carries a mutation in the active site. Besides the change in sugars in the cell walls, no other phenotypes are apparent. The mutant itself and knowledge of the gene product will be important for feedstock development. 2. Increasing starch in stems of switchgrass. A useful source of energy is starch, however, there is reluctance to trade potential food for biofuels. ARS scientists at Albany, CA, in collaboration with scientists as UC Berkeley have transformed the maize microRNA, Corngrass, into switchgrass. The plants have increased starch in their stems because the plants never flower. The lack of flowering is actually beneficial, as it prevents transgene seeds from escaping. Sugars can be obtained from these switchgrass plants without the use of costly pretreatments due to the increase in starch. Review Publications Moon, J., Hake, S.C. 2011. How a leaf gets its shape. Current Opinion in Plant Biology. 14:24-30.