Investigating the unusually high cell wall digestibility of the old INRA early flint F4 maize inbred line

The old INRA flint early line F4, which belongs to the northern flint group, is typified by its high cell wall digest- ibility which reaches values as high as those observed in several brown-midrib bm3 mutant lines. The F4 line thus appeared as a model that could contribute to the understanding of genetic mechanisms involved in variation of secondary wall traits. Different strategies and results were thus gathered including especially cell wall biochemical and digestibility investigations, expression approaches, QTL investigations, and colocalizations between QTLs and candidate genes. Lignin content was lower in F4 than in other lines, with a tendency to lower p-coumarate content. The Syringyl/Guaiacyl lignin unit ratio was similar in F4 as in other lines, but this ratio was nearly not reduced in F4bm3, conversely to what is observed in bm3 mutants. In comparison with the INRA F2 control line, expressions of three PAL genes including the ZmPAL, of the ZmF5H1 and the ZmCOMT genes were significantly reduced in F4 lignifying ear internodes at early silking stage. In the F7025 x F4 RIL progeny, seven QTLs were shown with favorable alleles (increasing cell wall digestibility) originating from F4. Two strong QTLs were located in bins 1.03 and 2.03 colocalizing with the ZmMYB019 and ZmSWN6 transcription factors, respectively. Orthologs of ZmMYB019 have been shown to be involved in lignin biosynthesis, and the PpMYB8 ortholog was shown to regu- late PAL gene expression in maritime pine. The ZmSWN6 NAC transcription factor is an upstream master regulator of the secondary wall biosynthetic programs. At the other QTL positions, colocalizations were also shown with other secondary wall related ZmMYB, but also with BAHD genes involved in arabinoxylan feruloylation, and with the position of the bm6 mutation. Three QTL positions were shown with favorable alleles originating from F7025, which colocalized with ZmMYB and ZmNAC transcriptions factors. As a tentative conclusion, the F4 unusually high cell wall digestibility is likely greatly related to the altered working of at least two major transcription factors regulating cell wall biosynthesis and assembly