Talalaiev O. Expression of small heat shock proteins in Pisum sativum L. seedlings in altered gravity

The thesis is addressed to the question of gravity-dependent gene expression in Pisum sativum. To determine the possible contribution of molecular chaperone genes in adaptation to the altered gravity conditions, we investigated gene expression of small heat shock proteins (sHsp) in etiolated pea seedlings under slow clinorotation (2 rpm) and centrifugation (3, 7, 10 та 14g). The sHsps are very diverse and variable in plants. Some higher plants have more than 30 individual sHsps genes and, unlike other groups, possess distinct sHsp subfamilies. Most of sHsp genes are highly sensitive to heat and other stressors. Others are selectively expressed in seeds and pollen, and a few are constitutively expressed. As a family, sHsp have a definite role in heat shock adaptation, but attributing specific effects to individual proteins has proved challenging. Methods of protein electrophoresis, immunodetection (Western blot) and RT-qPCR were used. Expression of five sHsp-genes belonging to the subfamilies with different subcellular localization: cytosolic-nuclear - Pshsp17.1 (class CII) and Pshsp18.1 (class-CI), chloroplast- Pshsp26.2 (class-P), mitochondrial - Pshsp22.9 (class-M) and endoplasmic reticulum - Pshsp22.7 (class-ER) has been examined. Expression of housekeeping gene - Actin was used for normalization of data. Based on qPCR results, we demonstrate that transcription of sHsp genes in 5-day old etiolated pea seedlings dramatically increases under 30?C and reached its maximum at 42?C. These results confirm sHsp genes as temperature sensitive and special role of sHsp under heat shock conditions. More sensitive to the temperature elevation were sHsp genes, coding mitochondrial, chloroplast and ER-localized small heat proteins. Relative qPCR results demonstrate that altered gravity and temperature elevations have different effects on the sHsp: unlike high temperature, altered gravity does not led to denaturation of cell proteins and, therefore, does not modulate sHsp gene expression.