Thenaked endospermGenes Encode Duplicate INDETERMINATE Domain Transcription Factors Required for Maize Endosperm Cell Patterning and Differentiation  

Abstract

The aleurone is the outermost layer of cereal endosperm and functions to digest storage products accumulated in starchy endosperm cells as well as to confer important dietary health benefits. Whereas normal maize (Zea mays [Zm]) has a single aleurone layer, naked endosperm (nkd) mutants produce multiple outer cell layers of partially differentiated cells that show sporadic expression of aleurone identity markers such as a viviparous1 promoter-b-glucuronidase transgene. The 15:1 F2 segregation ratio suggested that two recessive genes were involved, and map-based cloning identified two homologous genes in duplicated regions of the genome. The nkd1 and nkd2 genes encode the INDETERMINATE1 domain (IDD) containing transcription factors ZmIDDveg9 and ZmIDD9 on chromosomes 2 and 10, respectively. Independent mutant alleles of nkd1 and nkd2, as well as nkd2-RNA interference lines in which both nkd genes were knocked down, also showed the nkd mutant phenotype, confirming the gene identities. In wild-type kernels, the nkd transcripts were most abundant around 11 to 16 d after pollination. The NKD proteins have putative nuclear localization signals, and green fluorescent protein fusion proteins showed nuclear localization. The mutant phenotype and gene identities suggest that NKD controls a gene regulatory network involved in aleurone cell fate specification and cell differentiation.

NKD Transcription Factors Are Central Regulators of Maize Endosperm Development

“…Micrography was performed with a Jenoptik C-5 camera and constant gain 760 and exposure time settings were used for each filter set to compare expression of each 761 respective transgene reporter protein in WT versus nkd1 nkd2 mutant kernels. Standard 762 PCR genotyping of transgene and nkd1-R and nkd2-R alleles was performed using 763 primers described (Yi et al, 2015) to confirm kernel genotypes. 764…”

“…Kernel genotypes were 791 verified by standard PCR genotyping using primers described in Yi et al, 2015. Mature 792 kernels were cracked and freshly planed with a razor blade, cleaned with ethanol, and 793 placed on a specimen stub with a carbon coated adhesive. Specimen stubs were then 794 painted with silver paint and air dried for 10 minutes at room temperature.…”

“…47 4 To identify genes and biological processes directly or indirectly regulated by NKD1 and 91 NKD2 (NKD1/2) in developing endosperm, a transcriptomic analysis was undertaken to 92 identify gene transcripts differentially expressed (DE) between WT and nkd1 nkd2 93 mutant. Laser capture microdissection (LCM) coupled with RNAseq was previously 94 performed on AL and SE cells from WT (B73 inbred) versus nkd1 nkd2 mutant 95 endosperms at 15 days after pollination (DAP) (Yi et al, 2015). These 50-base paired-96 end reads were re-aligned and mapped to the maize reference genome (B73 RefGen-97 V3, GSE61057).…”

OPAQUE11 Is a Central Hub of the Regulatory Network for Maize Endosperm Development and Nutrient Metabolism

“…57 The AL is a single peripheral layer of cells that produces hydrolytic enzymes to 58 mobilize storage in the SE when the seed begins germination. The SE stores (Yi et al, 2015). Gontarek et al (2016) maturation and is also a repressor of germination-specific genes (McCarty et 77 al., 1991;Hoecker et al, 1995).…”

Mutations in the maize zeta-carotene desaturase gene lead to viviparous kernel

“…The maize y9 mutant has a slightly viviparous phenotype, and Y9 encodes a factor required for isomerase activity upstream of CRTISO, referred to as Z-ISO, and it has an activity that catalyzes the cis- to trans-conversion of the 15-cis-bond in 9, 15, 9’-tri-cis-zeta-carotene, the substrate of ZDS [20]. Apart from these typical viviparous mutants, the naked endosperm ( nkd ) mutants, which produce multiple outer cell layers, are prone to vivipary [21]. Luo and Wurtzel [22] isolated the cDNA of maize ZDS in 1999 and subsequently mapped this gene to the maize 7S chromosome.…”