Late embryogenesis abundant (LEA) proteins are large groups of hydrophilic proteins with major role in drought and other abiotic stresses tolerance in plants. In-depth study and characterization of LEA protein families have been carried out in other plants, but not in upland cotton. The main aim of this research work was to characterize the late embryogenesis abundant (LEA) protein families and to carry out gene expression analysis to determine their potential role in drought stress tolerance in upland cotton. Increased cotton production in the face of declining precipitation and availability of fresh water for agriculture use is the focus for breeders, cotton being the backbone of textile industries and a cash crop for many countries globally. In this work, a total of 242, 136 and 142 LEA genes were identified in G. hirsutum, G. arboreum and G. raimondii respectively. The identified genes were classified into eight groups based on their conserved domain and phylogenetic tree analysis. LEA 2 were the most abundant, this could be attributed to their hydrophobic character. Upland cotton LEA genes have fewer introns and are distributed in all chromosomes. Majority of the duplicated LEA genes were segmental. Syntenic analysis showed that greater percentages of LEA genes are conserved. Segmental gene duplication played a key role in the expansion of LEA genes. Sixty three miRNAs were found to target 89 genes, such as miR164, ghr-miR394 among others. Gene ontology analysis revealed that LEA genes are involved in desiccation and defense responses. Almost all the LEA genes in their promoters contained ABRE, MBS, W-Box and TAC-elements, functionally known to be involved in drought stress and other stress responses. Majority of the LEA genes were involved in secretory pathways. Expression profile analysis indicated that most of the LEA genes were highly expressed in drought tolerant cultivars Gossypium tomentosum as opposed to drought susceptible, G. hirsutum. The tolerant genotypes have a greater ability to modulate genes under drought stress than the more susceptible upland cotton cultivars. The finding provides comprehensive information on LEA genes in upland cotton, G. hirsutum and possible function in plants under drought stress.

Aquaporins (AQPs) are universal membrane integrated water channel proteins that selectively and reversibly facilitate the movement of water, gases, metalloids, and other small neutral solutes across cellular membranes in living organisms. Compared with other organisms, plants have the largest number of AQP members with diverse characteristics, subcellular localizations and substrate permeabilities. AQPs play important roles in plant water relations, cell turgor pressure maintenance, the hydraulic regulation of roots and leaves, and in leaf transpiration, root water uptake, and plant responses to multiple biotic and abiotic stresses. They are also required for plant growth and development. In this review, we comprehensively summarize the expression and roles of diverse AQPs in the growth and development of various vegetative and reproductive organs in plants. The functions of AQPs in the intracellular translocation of hydrogen peroxide are also discussed.

Abiotic stresses have negative effects on plants growth and development. Plants, being sessile, have developed specific adaptive strategies that allow them to rapidly detect and respond to abiotic stress factors. The detoxification efflux carriers (DTX)/multidrug and toxic compound extrusion (MATE) transporters are of significance in the translocation of abscisic acid (ABA), a phytohormone with profound role in plants under various abiotic stress conditions. The ABA signaling cascades are the core regulators of abiotic stress responses in plants, triggering major changes in gene expression and adaptive physiological responses. We therefore carried out genome-wide analysis of the DTX/MATE gene family, transformed a DTX/MATE gene in Arabidopsis and carried out functional analysis under drought, salt, and cold stress conditions. We identified 128, 70, and 72 DTX/MATE genes in Gossypium hirsutum, Gossypium arboreum, and Gossypium raimondii, respectively. The proteins encoded by the DTX/MATE genes showed varied physiochemical properties but they all were hydrophobic. The Gh_D06G0281 (DTX/MATE) over-expressing Arabidopsis lines were highly tolerant under drought, salt, and cold stress with high production of antioxidant enzymes and significantly reduced levels of oxidants. Lipid peroxidation, as measured by the level of malondialdehyde concentrations was relatively low in transgenic lines compared to wild types, an indication of reduced oxidative stress levels in the transgenic plants. Based on physiological measurements, the transgenic plants exhibited significantly higher relative leaf water content, reduced excised leaf water loss and a significant reduction in ion leakage as a measure of the cell membrane stability compared to the wild types. Abiotic stress responsive genes, ABF4, CBL1, SOS1, and RD29B were highly expressed in the transgenic lines compared to the non-transformed wild type plants. The protein encoded by the Gh_D06G0281 (DTX/MATE) gene was predicted to be located within the plasma membrane. Since signals from extracellular stimuli are transmitted through the plasma membrane most of which are conducted by plasma membrane proteins it is possible the Gh_D06G0281 (DTX/MATE) gene product could be important for this process.

We identified 102, 51 and 51 proteins encoded by the trihelix genes in Gossypium hirsutum, Gossypium arboreum and Gossypium raimondii, respectively. RNA sequence data and real‐time quantitative polymerase chain reaction analysis showed that Gh_A05G2067 (GT‐2) was highly upregulated under drought and salt stress conditions. Transient expression of GT‐2‐green fluorescent protein fusion protein in protoplast showed that GT‐2 was localized in the nucleus. The overexpression of GT‐2 conferred an enhanced drought tolerance to cotton, with lower malondialdehyde, hydrogen peroxide contents and higher reactive oxygen scavenging enzyme activities. Moreover, chlorophyll content, relative leaf water content (RLWC), excised leaf water loss (ELWL) and cell membrane stability (CMS) were relatively stable in the GT‐2‐overexpressed lines compared to wild‐type (WT). Similarly, stress‐responsive genes RD29A, SOS1, ABF4 and CBL1 were highly upregulated in the GT‐2‐overexpressed lines but were significantly downregulated in WT. In addition, the GT‐2‐silenced cotton plants exhibited a high level of oxidation injury, due to high levels of oxidant enzymes, in addition to negative effects on CMS, ELWL, RLWC and chlorophyll content. These results mark the foundation for future exploration of the trihelix genes in cotton, with an aim of developing more resilient, versatile and highly tolerant cotton genotypes.

Increasing soil salinization has led to severe reductions in plant yield and quality, and investigating the molecular mechanism of salt stress response is therefore an urgent priority. In this study, we systematically analyzed the response of cotton roots to salt stress using single-cell transcriptomics technology; 56 281 high-quality cells were obtained from 5-day-old lateral root tips of Gossypium arboreum under natural growth conditions and different salt treatments. Ten cell types with an array of novel marker genes were identified and confirmed by in situ RNA hybridization, and pseudotime analysis of some specific cell types revealed their potential differentiation trajectories. Prominent changes in cell numbers under salt stress were observed for outer epidermal and inner endodermal cells, which were significantly enriched in response to stress, amide biosynthetic process, glutathione metabolism, and glycolysis/gluconeogenesis. Analysis of differentially expressed genes identified in multiple comparisons revealed other functional aggregations concentrated on plant-type primary cell wall biogenesis, defense response, phenylpropanoid biosynthesis, and metabolic pathways. Some candidate differentially expressed genes encoding transcription factors or associated with plant hormones also responsive to salt stress were identified, and the function of Ga03G2153, annotated as auxin-responsive GH3.6, was confirmed by virus-induced gene silencing. The GaGH3.6-silenced plants showed a severe stress-susceptible phenotype, and physiological and biochemical measurements indicated that they suffered more significant oxidative damage. These results suggest that GaGH3.6 might participate in cotton salt tolerance by regulating redox processes. We thus construct a transcriptional atlas of salt-stressed cotton roots at single-cell resolution, enabling us to explore cellular heterogeneity and differentiation trajectories and providing valuable insights into the molecular mechanisms that underlie plant stress tolerance.

The extrusion of toxins and substances at a cellular level is a vital life process in plants under abiotic stress. The multidrug and toxic compound extrusion (MATE) gene family plays a large role in the exportation of toxins and other substrates. We carried out a genome-wide analysis of MATE gene families in Gossypium raimondii and Gossypium arboreum and assessed their expression levels under salt, cadmium and drought stresses. We identified 70 and 68 MATE genes in G. raimondii and G. arboreum, respectively. The majority of the genes were predicted to be localized within the plasma membrane, with some distributed in other cell parts. Based on phylogenetic analysis, the genes were subdivided into three subfamilies, designated as M1, M2 and M3. Closely related members shared similar gene structures, and thus were highly conserved in nature and have mainly evolved through purifying selection. The genes were distributed in all chromosomes. Twenty-nine gene duplication events were detected, with segmental being the dominant type. GO annotation revealed a link to salt, drought and cadmium stresses. The genes exhibited differential expression, with GrMATE18, GrMATE34, GaMATE41 and GaMATE51 significantly upregulated under drought, salt and cadmium stress, and these could possibly be the candidate genes. Our results provide the first data on the genome-wide and functional characterization of MATE genes in diploid cotton, and are important for breeders of more stress-tolerant cotton genotypes.