Exploring Wheat Grain Nutrient Content, Nutrient Remobilisation and Senescence Using the NAM-B1 Transcription Factor.

NAM-B1 is a NAC transcription factor which is an important regulator of wheat grain nutrient content and monocarpic senescence. NAM-B1 was originally cloned as the gene underlying the Gpc-B1 QTL which increases grain protein content. It was also found that NAM-B1 affects grain micronutrient content, particularly iron and zinc. These changes to grain nutrient content are related to a delay in flag leaf senescence, suggesting that NAM-B1 controls remobilisation of nutrients from this leaf to the developing grain. We are investigating the function of NAM-B1 by identifying its direct targets using ChIP-seq and by physiological and molecular characterisation.

To understand how NAM-B1 controls grain nutrient content, nutrient remobilisation and senescence we have identified direct targets regulated by NAM-B1 using ChIP-seq (Chromatin Immuno-Precipitation combined with next-generation sequencing). We have identified several hundred potential binding sites of NAM-B1 which are mainly found within 1kb of the open reading frame of the nearest gene, suggesting a regulatory role for the identified binding sites. To confirm the regulatory effect of NAM-B1 on these identified genes we are using RNA-seq in wheat plants with reduced expression of NAM-B1 to identify genes which are both directly bound by NAM-B1 and differentially expressed knock-down plants. We will identify TILLING mutants in a subset of these high-confidence candidate genes to further dissect the pathways of nutrient remobilisation, senescence and grain nutrient content.

In parallel we have examined the expression patterns of NAM-B1 finding that it is expressed in stem, peduncle, leaf and grain tissues after anthesis. Using RNA in situ hybridisation we found that NAM-B1 is expressed in the vascular tissues and we hypothesised it might regulate xylem and phloem transport of nutrients to the grain.  We carried out a tracer experiment and found that xylem and phloem transport are unaltered in plants with reduced expression of NAM-B1, leading us to conclude that NAM-B1 did not alter nutrient transport at the level of xylem and phloem.