Abstract
The black layer (BL) is traditionally used as an indicator for kernel harvesting in maize, as it turns visibly dark when the kernel reaches physiological maturity. However, the molecular roles of BL in kernel development have not been fully elucidated. In this work, microscopy images showed that BL began to appear at a growth stage earlier than 10 days after pollination (DAP), and its color gradually deepened to become dark as the development period progressed. Scanning electron microscopy observations revealed that BL is a tissue structure composed of several layers of cells that are gradually squeezed and compressed during kernel development. Laser-capture microdissection (LCM) was used to sample BL and its neighboring inner tissue, basal endosperm transfer layer (BETL), and outer tissue, inner epidermis (IEP), from 20 DAP of kernels. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry profiling (MALDI-TOF MS profiling) detected 41, 104, and 120 proteins from LCM-sampled BL, BETL, and IEP, respectively. Gene ontology (GO) analysis indicated that the 41 BL proteins were primarily involved in the response to stress and stimuli. Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway analysis found that the BL proteins were enriched in several defense pathways, such as the ascorbate and aldarate metabolic pathways. Among the 41 BL proteins, six were BL-specific proteins that were only detected from BL. Annotations of five BL-specific proteins were related to stress responses. During kernel development, transcriptional expression of most BL proteins showed an increase, followed by a decrease, and reached a maximum zero to 20 DAP. These results suggest a role for BL in stress responses for protecting filial tissue against threats from maternal sides, which helps to elucidate the biological functions of BL.
Highlights
Maize is one of the most important grain crops with a total production of more than one billion tons, accounting for ~30% of the world’s food supply
Black layer (BL) is derived from the inner integument and the integumental P-C, which is located immediately below another P-C layer, nucellar P-C which is immediately subtended from the endosperm and is derived from the nucellus epidermis [10]
At the early kernel development stage, programmed cell death process (PCD)-based cell death was observed in BL [11], which leads to cell degradation into a narrow band of crushed cells with accumulated brown pigment and subsequently leads to the emergence of visible BL [11,12]
Summary
Maize is one of the most important grain crops with a total production of more than one billion tons, accounting for ~30% of the world’s food supply (https://www.statista.com). At the early kernel development stage (before 10 days after pollination, DAP), programmed cell death process (PCD)-based cell death was observed in BL [11], which leads to cell degradation into a narrow band of crushed cells with accumulated brown pigment and subsequently leads to the emergence of visible BL [11,12]. These visible BLs are generally used as kernel maturity indicators by agronomists and farmers. These results could help to elucidate the mechanisms of BL formation and the biological roles of this special structure
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