Muhammad Shafiq

Background Carotenoid cleavage oxygenases (CCOs) are non-heme iron-dependent enzymes that catalyze the oxidative cleavage of carotenoids and play important roles in the biosynthesis of apocarotenoids involved in plant growth, development, and stress responses. However, a comprehensive genome-wide characterization of the CCO gene family in barley (Hordeum vulgare L.) has not been reported. Methods and Results In this study, a genome-wide analysis was performed to identify and characterize CCO genes in barley using comparative genomics and bioinformatics approaches. A total of 11 HvCCO genes were identified and classified into three subfamilies: carotenoid cleavage dioxygenases (CCDs), carotenoid cleavage dioxygenase-like proteins (CCDLs), and 9-cis-epoxycarotenoid dioxygenases (NCEDs). Gene structure and conserved motif analyses revealed high conservation within subfamilies, while promoter analysis identified 190 cis-regulatory elements associated with light, phytohormone, stress, and developmental regulation. Chromosomal mapping, duplication, and synteny analyses indicated that segmental duplication contributed to HvCCO gene family expansion, while Ka/Ks analysis suggested purifying selection of duplicated gene pairs. Prediction of miRNA targets identified 12 barley miRNAs potentially regulating HvCCO genes. Publicly available RNA-seq datasets revealed tissue-specific expression patterns. Under abiotic and biotic stresses, HvCCD4 and HvCCD6 showed relatively consistent expression, whereas HvNCED1 and HvNCED2 exhibited increased expression under drought conditions in tolerant genotypes. Conclusions These findings provide a comprehensive genomic resource for the barley CCO gene family and identify HvCCD4 and HvCCD6 as potential candidate genes for future functional validation and genetic improvement of stress tolerance in barley.

Pathogenesis-related (PR) proteins are critical components of the plant immune system, rapidly accumulating in response to pathogen attack and various stresses. Since their first identification in tobacco infected with Tobacco Mosaic Virus (TMV), these inducible proteins have been classified into multiple families based on sequence and functional characteristics. PR proteins exhibit a range of biochemical activities – including cell-wall degrading enzyme activities (e.g., chitinases, glucanases), antimicrobial peptide functions (e.g., defensins, thionins), and protease inhibition – that contribute to direct pathogen suppression and modulation of defense signaling. Advances in molecular cloning have enabled transgenic expression of PR genes in susceptible crops, often leading to enhanced resistance against fungi, bacteria, oomycetes, and viruses. Beyond plant immunity, some PR proteins display multifunctional properties relevant to stress tolerance and pharmacology. This review summarizes current understanding of PR protein classification, molecular features, induction by biotic/abiotic cues, and their application in crop protection, while highlighting opportunities and challenges in harnessing PR proteins for biotechnology.

In higher plants, YABBY transcription factors are critical for the development of lateral organs, polarity determination, and the response to environmental stress. Although the olive tree, Olea europaea L., is a valuable crop species in terms of ecology and economics, the YABBY gene family has received little attention. The current work discovered five unique OeuYAB genes from the five known subfamilies of the YABBY gene family: FIL/YAB3, YAB2, YAB5, CRC, and INO. Chromosomal mapping revealed that the distribution of these genes is not consistent throughout five genomic areas, denoted as SC_2771, SC_528, C_9, C_3, and C_1. Oeu055459.1 is placed at the distal end of C_1, while Oeu033104.1 and Oeu048822.1 are found in the higher sections of SC_2771 and SC_528, respectively. The clustering of Oeu055459.1, Oeu055459.2, and Oeu055459.3 in genomic regions C_9 and C_3 indicates the presence of genomic hotspots that could have resulted from the original gene's duplication. The physicochemical characterization of the genes revealed that OeuYAB proteins are compact (170-211 amino acids), hydrophilic, and unstable. The structural and motif analysis revealed that the genes contain the N-terminal C2C2 Zinc Finger and C-terminal YABBY domains. The Ka/Ks ratio is less than one, indicating strong purifying selection pressure on these genes. Furthermore, the presence of a complex regulatory system in the genes' promoters revealed that they are high in light responsiveness, phytohormones, and abiotic stress adaptation. To investigate post-transcriptional regulation, a miRNA target analysis revealed a specialized mode of regulation in which the OeuYAB1 and OeuYAB4 genes are regulated by two distinct miRNA clusters, Oeu-miRN1154 and Oeu-miRN2138, for translational inhibition and transcript cleavage, respectively. The OeuYAB genes were profiled using RNA-seq expression profiling in six anatomical tissues, revealing a high level of functional divergence; for example, OeuYAB3 of the CRC subfamily was found to have the highest expression level (1081.066 RPKM) in the fruit tissue, establishing its role as a key candidate for olive drupe development.

Pathogenesis-related gene PR1 acts as an essential element in the plant defensive mechanism and is made during the process of systemic acquired resistance (SAR) and salicylic-induced signaling. The genome-wide analysis of the PR1 gene in Pisum sativum has been conducted to analyze the functionality, composition, and evolutionary status of this gene. Based on the fact that CAP (Cysteine-rich secretory proteins, Antigen 5, and Pathogenesis-related 1) domain, fifteen PsPR1 genes were identified. Complete CAP domain has been found on these proteins, which ensures that all belong to the PR1 family. There is a mild difference in the size of protein, the molecular mass, as well as the point of isoelectric activity showed by the physicochemical study. On the other hand, subcellular localization predictions revealed that there is predominant extracellular targeting, with the secretory nature of PR-1 proteins, in multiple cellular compartments. The Fabaceae family has a lineage-specific increase of PR-1 genes, according to phylogenetic research with several dicot species. This suggests functional conservation with known Arabidopsis thaliana PR-1 homologs involved in SAR. The majority of PsPR1 genes are either intron-less or have a single intron, supporting fast transcriptional activation during stress, according to conserved motif and gene structure analyses that revealed strong evolutionary conservation. Strong purifying selection was shown by Ka/Ks analysis of duplicated gene pairs, highlighting PR-1 genes' physiological importance in pea defence. Numerous cis-regulatory regions linked to hormone signalling, stress response, and transcription factor binding were found by promoter analysis, suggesting intricate regulatory control. PsPR1 genes were found to be distributed unevenly by chromosomal mapping, with tandem duplication being the main cause of the substantial clustering on chromosome 1. Tandem and segmental duplication events both contributed to the formation of the gene family, according to synteny analysis. PsPR1.1, PsPR1.3, and PsPR1.9 were found to be significantly regulated genes by expression profiling during heat stress, especially in the heat-tolerant cultivar Akra Chaitra. Overall, this study identifies intriguing possibilities for enhancing heat stress resilience and offers a solid foundation for comprehending the functional evolution and stress-responsive activities of PR-1 genes in Pisum sativum.

Abiotic stressors like drought, salinity, heat, heavy metals, oxidative stress, and UV radiation severely limit plant growth, development, and productivity. Plants detect these stresses using particular membrane sensors, resulting in fast changes in intracellular calcium concentrations, reactive oxygen species pulses, and phytohormone signaling. These initial signals are converted into conserved kinase signaling pathways (MAPKs, CDPKs, SnRK2s) and hormone-mediated pathways (ABA-dependent and ABA-independent), which then activate stress-responsive transcription factors (DREBs, NACs, WRKYs, MYBs) and remodel the transcriptome. Chromatin modification, alternative splicing, short RNAs, and post-translational modifications (phosphorylation and ubiquitination) all contribute to complicated regulation, ensuring precise control of stress gene expression. Plants use hormonal interactions and network hubs to balance survival, growth, and defense. Recent advances in systems biology have revealed these complicated networks, and biotechnological approaches—transgenic methods, CRISPR/Cas genome editing, and multi-omics integration—have opened up new avenues for the production of stress-tolerant crops. This chapter provides a thorough, human-crafted overview of these processes and examines future directions for applying molecular knowledge to sustainable farming operations.

Climate change offers a growing danger to global horticulture productivity, with rising temperatures, shifting precipitation patterns, and increased soil salinity. However, throughout the last decade, the GWAS era has revolutionized our understanding of genetic architecture for abiotic stress tolerance in horticulture crops. This study outlines GWAS research development from 2015 to 2025, focusing on drought, heat, and salinity tolerance in horticulture crops. Some of the significant highlights are the finding of stable loci such as SlALMT15 in tomato for stomatal function, QTL for tuber heat tolerance in potato, and salt-tolerant genes in cucumber seedlings. Integration with transcriptomics, phenomics, and genomic selection has accelerated candidate gene validation and breeding applications. Despite these advancements, critical challenges remain—limited sample sizes, inconsistent phenotyping, underrepresentation of orphan crops, and the need for multi-environment trials. Abiotic stressors account for 50-80% of the potential yield loss of numerous horticulture crops, according to global studies conducted between 2010 and 2025. The combination of multi-omics and AI-assisted genomic prediction has increased the effectiveness of GWAS-based trait identification in plant breeding research. This review also explores the role of international consortia, community genomics databases, and data-sharing protocols in enhancing GWAS effectiveness. By aligning genomics, phenotyping, and breeding, GWAS serves as a transformative approach for developing climate-smart horticultural cultivars. Strengthening global collaboration, expanding diversity panels, and applying advanced analytics are key to building future-resilient food systems.

Drought stress imposes a critical limitation on sugarcane productivity, thereby necessitating the development of stress-resilient cultivars for sustainable agriculture. The present study aimed to enhance drought tolerance in sugarcane by introducing the Arabidopsis DREB1A gene under the control of the stress-inducible rd29A promoter. Transgenic sugarcane lines were generated via particle bombardment-mediated transformation and subsequently evaluated through molecular, physiological, and agronomic assessments under controlled and drought-stressed conditions. Molecular analysis confirmed stable transgene integration, with transgenic lines exhibiting up to a 10-fold increase in DREB1A expression relative to wild-type plants. Physiological assessments demonstrated that, under drought stress at 60% field capacity (FC), transgenic lines maintained photosynthetic rates (PN) that were 224-270% higher, stomatal conductance (gs) increased by 84-167%, and relative water content (RWC) was enhanced by 25-31% compared to non-transgenic controls. Moreover, the leaves of transgenic sugarcane displayed improved osmotic regulation and water-use efficiency. Agronomic evaluations further revealed significant improvements in plant growth and productivity. Under drought stress (60% FC), transgenic lines exhibited 76-109% greater cane height, with cane diameter 71-86% larger. Shoot biomass increased by 39-87%, and root biomass was enhanced by 65-103%. Additionally, the Brix percentage, indicative of sucrose accumulation, increased by 36-55% in the transgenic plants at 60% FC. These findings establish a robust correlation between DREB1A expression, enhanced physiological resilience, and improved agronomic performance under drought conditions. The capacity of DREB1A-expressing transgenic sugarcane to sustain higher photosynthetic activity, superior water-use efficiency, and increased biomass accumulation underscores its potential as a genetic strategy for developing drought-resilient sugarcane varieties. This study offers novel insights into the molecular mechanisms underlying drought tolerance and provides a promising approach for ensuring sustainable sugarcane cultivation in water-scarce regions.

Drought stress imposes a critical limitation on sugarcane productivity, thereby necessitating the development of stress-resilient cultivars for sustainable agriculture. The present study aimed to enhance drought tolerance in sugarcane by introducing the Arabidopsis DREB1A gene under the control of the stress-inducible rd29A promoter. Transgenic sugarcane lines were generated via particle bombardment-mediated transformation and subsequently evaluated through molecular, physiological, and agronomic assessments under controlled and drought-stressed conditions. Molecular analysis confirmed stable transgene integration, with transgenic lines exhibiting up to a 10-fold increase in DREB1A expression relative to wild-type plants. Physiological assessments demonstrated that, under drought stress at 60% field capacity (FC), transgenic lines maintained photosynthetic rates (PN) that were 224-270% higher, stomatal conductance (gs) increased by 84-167%, and relative water content (RWC) was enhanced by 25-31% compared to non-transgenic controls. Moreover, the leaves of transgenic sugarcane displayed improved osmotic regulation and water-use efficiency. Agronomic evaluations further revealed significant improvements in plant growth and productivity. Under drought stress (60% FC), transgenic lines exhibited 76-109% greater cane height, with cane diameter 71-86% larger. Shoot biomass increased by 39-87%, and root biomass was enhanced by 65-103%. Additionally, the Brix percentage, indicative of sucrose accumulation, increased by 36-55% in the transgenic plants at 60% FC. These findings establish a robust correlation between DREB1A expression, enhanced physiological resilience, and improved agronomic performance under drought conditions. The capacity of DREB1A-expressing transgenic sugarcane to sustain higher photosynthetic activity, superior water-use efficiency, and increased biomass accumulation underscores its potential as a genetic strategy for developing drought-resilient sugarcane varieties. This study offers novel insights into the molecular mechanisms underlying drought tolerance and provides a promising approach for ensuring sustainable sugarcane cultivation in water-scarce regions.