Research Objective
To investigate heat stress effects on ovule fertility and ovule damage in ovaries of pea; to measure heat stress effects on ovule fertility and damage in ovaries for 18 genotypes of pea.
Pea yield is sensitive to heat stress, and the crop loses pods and seeds in warm years. Poor pollen and pollen viability are part of the problem as heat damages pollination, but little is known about the heat robustness or fate of ovules in heat stress. Ovules, once fertilized by pollen, are destined to become pea seeds in pods. Prior to this project, we believed that pea varieties and their ovules in pods have a range of heat sensitivity to resistance. Some varieties are more likely to keep all their seeds in pods in warm summers, and we must determine these pea varieties.
Pea yield is sensitive to heat stress and the crop loses pods and seeds in warm years. Heat damages pollination processes during flowering, but little is known about ovules in heat stress. Ovules, after being fertilized by pollen, are destined to become pea seeds within the ovary or pea pod. Pea varieties and their ovules have a range of heat sensitivity to resistance because some varieties keep most of their seeds but other varieties lose many under heat conditions. The overall goal of this research was to complement physical measurements taken with light microscopy and investigate if pea ovules had programmed cell death, and to look at the change in gene expression in young ovules that had experienced heat.
To do this we grew three varieties that were heat sensitive (Carneval), somewhat heat robust (CDC Meadow), and heat robust (Naparnyk) in growth chambers under control conditions (normal temperatures) and then treated half of them at 35 degrees Celsius for four days at the start of flowering. We sampled the pods at the second flowering node, which would have just been pollinated on the day we started the heat treatment. After four days, we measured the ovules within pods for the effects of heat stress using a microscope staining method that uses fluorescence to indicate programmed cell death. This method is called TUNEL staining, and it stains degraded DNA. The second method looks at gene expression using RNA and can measure which genes are turned on more (upregulated) or turned down more (down regulated). The RNA method allowed us to look at gene expression in ovules across the range of the three varieties, and under normal (control) versus heat treatments.
Our preliminary results show that pea ovules from node two of Carneval, the most heat susceptible variety we measured, do not undergo widespread programmed cell death, and only a few small areas in ovules were affected by heat and then only slightly. In Carneval, the top downregulated genes in heat were stress responsive genes, particularly those with hydrolase activities. In contrast, genes of that group were up-regulated in Naparynk, the heat robust cultivar. So Naparnyk has a better stress response to heat and this cultivar turns up the signal through its stress responsive genes. Naparnyk’s most down regulated genes were ones that modified the cell wall. These results lead us closer to candidate genes for heat resistance, so we can screen for heat resistance easily, automate our methods, and produce improved pea varieties.