Omics approaches to understand the MADS-box gene family in common bean (Phaseolus vulgaris L.) against drought stress.

MADS-box genes are known to play important roles in diverse aspects of growth/devolopment and stress response in several plant species. However, no study has yet examined about MADS-box genes in P. vulgaris. In this study, a total of 79 PvMADS genes were identified and classified as type I and type II according to the phylogenetic analysis. While both type I and type II PvMADS classes were found to contain the MADS domain, the K domain was found to be present only in type II PvMADS proteins, in agreement with the literature. All chromosomes of the common bean were discovered to contain PvMADS genes and 17 paralogous gene pairs were identified. Only two of them were tandemly duplicated gene pairs (PvMADS-19/PvMADS-23 and PvMADS-20/PvMADS-24), and the remaining 15 paralogous gene pairs were segmentally duplicated genes. These duplications were found to play an important role in the expansion of type II PvMADS genes. Moreover, the RNAseq and RT-qPCR analyses showed the importance of PvMADS genes in response to drought stress in P. vulgaris.

Characterization of Two-Component System gene (TCS) in melatonin-treated common bean under salt and drought stress.

The two-component system (TCS) generally consists of three elements, namely the histidine kinase (HK), response regulator (RR), and histidine phosphotransfer (HP) gene families. This study aimed to assess the expression of TCS genes in P. vulgaris leaf tissue under salt and drought stress and perform a genome-wide analysis of TCS gene family members using bioinformatics methods. This study identified 67 PvTCS genes, including 10 PvHP, 38 PvRR, and 19 PvHK, in the bean genome. PvHK2 had the maximum number of amino acids with 1261, whilst PvHP8 had the lowest number with 87. In addition, their theoretical isoelectric points were between 4.56 (PvHP8) and 9.15 (PvPRR10). The majority of PvTCS genes are unstable. Phylogenetic analysis of TCS genes in A. thaliana, G. max, and bean found that PvTCS genes had close phylogenetic relationships with the genes of other plants. Segmental and tandem duplicate gene pairs were detected among the TCS genes and TCS genes have been subjected to purifying selection pressure in the evolutionary process. Furthermore, the TCS gene family, which has an important role in abiotic stress and hormonal responses in plants, was characterized for the first time in beans, and its expression of TCS genes in bean leaves under salt and drought stress was established using RNAseq and qRT-PCR analyses. The findings of this study will aid future functional and genomic studies by providing essential information about the members of the TCS gene family in beans. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-023-01406-5.

The genome-wide characterization of WOX gene family in Phaseolus vulgaris L. during salt stress.

The WUSCHEL-Related Homeobox (WOX) family is a type of homeobox transcription factor superfamily and its members perform many functions ranging from plant embryonic growth to organ formation in plants. Although the WOX proteins have been identified and characterized in many plant species, genome-wide identification and characterization of WOX proteins in the Phaseolus vulgaris genome has been performed for the first time in this study. Accordingly, 18 WOX proteins were identified using bioinformatics tools and biochemical/physicochemical properties of these proteins were investigated. Phvul-WOX genes were found to be categorized into three major phylogenetic groups according to the phylogenetic analysis and a total of five segmental duplication events were detected after duplication analysis. Moreover, the Phvul-WOX genes were found to be expressed in different plant tissues at different levels and some stress-related miRNAs have been found to target the Phvul-WOX genes based on miRNA analysis. Additionaly, MDA content, total protein level and catalase enzyme activity analyses were conducted in two P. vulgaris cultivars namely Yakutiye cv. and Zulbiye cv. subjected to 150 mM salt stress. Next, these cultivars were used for screening the expression levels of Phvul-WOX-1, Phvul-WOX-9, Phvul-WOX-11, Phvul-WOX-15 and Phvul-WOX-16 genes in response to salt stress. The insights gained from this study may be of assistance to the researchers who work in this area. Supplementary Information: The online version contains supplementary material available at 10.1007/s12298-022-01208-1.

Identification and Characterization of SRS Genes in Phaseolus vulgaris Genome and Their Responses Under Salt Stress.

SHI-Related Sequence (SRS) transcription factors comprise a protein family with important roles in growth and development. However, the genome-wide study of the SRS protein family has not yet been carried out in the common bean. For this reason, the SRS family has been characterized in depth at both gene and protein levels and several bioinformatics methods have been used. As a result, 10 SRS genes have been identified and their proteins have been phylogenetically categorized into three major groups within the common bean. By investigating duplications that play a major role in the development of gene families, 19 duplication events have been identified in the SRS family (18 segmental and 1 tandem). In addition, using available RNAseq data, comparative expression analysis of Pvul-SRS genes was performed and expression changes in Pvul-SRS-1, 2, 4, 6, 7, and 10 genes were observed under both salt and drought stress. Five Pvul-SRS genes were selected based on RNAseq data (Pvul-SRS-1, 2, 4, 6, and 10) and screened with RT-qPCR in two common bean cultivars (Yakutiye 'salt-resistant' and Zulbiye 'salt-susceptible' cv.). These genes also showed different levels of expression between two common bean cultivars under salt stress conditions and this may explain the responses of Pvul-SRS genes against abiotic stress. In summary, this work is the first study in which in silico identification and characterization of Pvul-SRS genes have been examined at gene expression level. The results could therefore provide the basis for future studies of functional characterization of Pvul-SRS genes.

Eastern Anatolian apples with a unique population structure are genetically different from Anatolian apples.

The origin of the apple is known to be the Transcaucasian region. Eastern Anatolia, which is located on the migration routes from Asia to Europe, has a rich and an uncharacterized apple germplasm and the characterization of apple genetic sources from this region is important for both evolutionary studies and apple breeding. In this study, 94 M. domestica accessions originated from seven diverse regions within Eastern Anatolia were studied using 16 SSR (simple sequence repeat) loci. SSR markers we used produced high allele numbers in all loci and CH02d11 (PI: 0.059) with 18 alleles was the most informative locus. In addition, 14 identical accession groups were identified. Most likely due to self-incompatibility, relatively high levels of heterozygosity (Ho: 0.696) was found for Eastern Anatolia apples. Structure Harvester analyses of East Anatolian apple accessions showed that although each group seems to be somewhat distinct, some levels of admixture with other populations might also exist. Due to a significant gene flow between all pairs of seven apple populations, a limited (low) differentiation was found between the populations. Comparisons using 16 common SSR loci revealed that Eastern Anatolia accessions were genetically different from Anatolian accessions. In addition, based on FCA, and Nei's genetic distance analyses, Eastern Anatolian apples were found to be genetically different from the commercial apple cultivars Golden Delicious and Florina. Our results suggesting that Eastern Anatolia apple populations have a unique structure will be useful for future genetic and evolutionary studies on apples.

Genome-wide identification of CAMTA gene family members in Phaseolus vulgaris L. and their expression profiling during salt stress.

The calmodulin-binding transcriptional activator (CAMTA) family was first observed in tobacco (NtER1) during a screening for the CaM-binding proteins, which are known to be one of the fast response stress proteins. Due to the increased importance of plant transcription factors in recent years; genome-wide identification of CAMTA genes has been performed in several plant species, except for Phaseolus vulgaris. Therefore, our aim was to identify and characterize CAMTA genes in P. vulgaris via in silico genome-wide analysis approach. Our results showed a total of eight CAMTA genes that were identified and observed on five out of 11 chromosomes of P. vulgaris. Four gene couples were found to be segmentally-duplicated and these segmental duplication events were shown to occur from 29.97 to 92.06 MYA. The phylogenetic tree of CAMTA homologs from P. vulgaris, A. thaliana, and G. max. revealed three groups based on their homology and the intron numbers of Pvul-CAMTA genes, ranged from 11 to 12. According to the syteny analysis; CAMTA genes of P. vulgaris and G. max revealed higher similarity, because they have highly similar genomes compared to A. thaliana. All Pvul-CAMTA genes were targeted by miRNAs, which play a role in response mechanism of salt stress. To detect expression levels in different plant tissues, mRNA analysis of Pvul-CAMTA genes were performed using publicly available expression data in Phytozome v12.1. In addition, responses of Pvul-CAMTA genes to salt stress, were also examined via both RNAseq and qRT-PCR analysis. To identify and to obtain insight into biological functions of CAMTA genes in the genome of P. vulgaris, several analyses were conducted using many online and offline bioinformatic tools, genome databases and qRT-PCR analyses. Due to this study being the first in the identification of CAMTA genes in P. vulgaris, this study could be considered as an useful source for future CAMTA genes studies in either P. vulgaris or comparative different plant species.

Determination of Usnic Acid Responsive miRNAs in Breast Cancer Cell Lines.

OBJECTIVE: Breast Cancer (BC) is the most common type of cancer diagnosed in women. A common treatment strategy for BC is still not available because of its molecular heterogeneity and resistance is developed in most of the patients through the course of treatment. Therefore, alternative medicine resources as being novel treatment options are needed to be used for the treatment of BC. Usnic Acid (UA) that is one of the secondary metabolites of lichens used for different purposes in the field of medicine and its anti-proliferative effect has been shown in certain cancer types, suggesting its potential use for the treatment. METHODS: Anti-proliferative effect of UA in BC cells (MDA-MB-231, MCF-7, BT-474) was identified through MTT analysis. Microarray analysis was performed in cells treated with the effective concentration of UA and UA-responsive miRNAs were detected. Their targets and the pathways that they involve were determined using a miRNA target prediction tool. RESULTS: Microarray experiments showed that 67 miRNAs were specifically responsive to UA in MDA-MB-231 cells while 15 and 8 were specific to BT-474 and MCF-7 cells, respectively. The miRNA targets were mostly found to play role in Hedgehog signaling pathway. TGF-Beta, MAPK and apoptosis pathways were also the prominent ones according to the miRNA enrichment analysis. CONCLUSION: The current study is important as being the first study in the literature which aimed to explore the UA related miRNAs, their targets and molecular pathways that may have roles in the BC. The results of pathway enrichment analysis and anti-proliferative effects of UA support the idea that UA might be used as a potential alternative therapeutic agent for BC treatment.

Transcriptome - Scale characterization of salt responsive bean TCP transcription factors.

TEOSINTE-BRANCHED1/CYCLOIDEA/PCF (TCP) proteins are important regulators of growth and developmental processes including branching, floral organ morphogenesis and leaf growth as well as stress response. This study identified 27 TCP genes of Phaseolus vulgaris (common bean), which were divided into three clusters based on phylogenetic relationship. In addition, this study showed that some of TCP genes such as Pvul-TCP-4 and Pvul-TCP-15 located on chromosomes 3 and 7, Pvul-TCP-7 and Pvul-TCP-20 located on chromosome 7 and 9, were segmentally duplicated. On the other hand, a total of 20 Pvul-TCP genes have predicted to be targeted by microRNAs (miRNA). Most of the miRNA-target genes were Pvul-TCP-1, -11, -13 and -27, which were targeted by 13, 17, 22 and 13 plant miRNAs, respectively. miR319 was one of the highly represented regulatory miRNAs to target TCP transcripts. Promoter region analysis of TCP genes resulted that the GT-1 motif, which was related to salt stress, was found in 14 different Pvul-TCP genes. Expression profiling of 10 Pvul-TCP genes based on RNA-sequencing data further confirmed with quantitative real-time RT-PCR measurements identified that Pvul-TCP genes under salt stress are expressed in a cultivar- and tissue-specific manner.

Genome-wide analysis of Phaseolus vulgaris C2C2-YABBY transcription factors under salt stress conditions.

The aim of this study was to identify and characterize the C2C2-YABBY family of genes by a genome-wide scale in common bean. Various in silico approaches were used for the study and the results were confirmed through common molecular biology techniques. Quantitative real-time PCR (qPCR) analysis was performed for identified putative PvulYABBY genes in leaf and root tissues of two common bean cultivars, namely Yakutiye and Zulbiye under salt stress condition. Eight candidate PvulYABBY proteins were discovered and the length of these proteins ranged from 173 to 256 amino acids. The isoelectric points (pIs) of YABBY proteins were between 5.18 and 9.34 and ranged from acidic to alkaline, and the molecular weight of PvulYABBYs were between 18978.4 and 28916.8 Da. Three segmentally duplicated gene couples among the identified eight PvulYABBY genes were detected. These segmentally duplicated gene couples were PvulYABBY-1/PvulYABBY-3, PvulYABBY-5/PvulYABBY-7 and PvulYABBY-6/PvulYABBY-8. The predicted number of exons among the PvulYABBY genes varied from 6 to 8 exons. Additionally, all genes found included introns within ORFs. PvulYABBY-2, -4, -5 and -7 genes were targeted by miRNAs of five plant species and a total of five miRNA families (miR5660, miR1157, miR5769, miR5286 and miR8120) were detected. According to RNA-seq analysis, all genes were up- or down-regulated except for PvulYABBY-1 and PvulYABBY-6 after salt stress treatment in leaf and root tissues of common bean. According to the qPCR analysis, six out of eight genes were expressed in the leaves but only four out of eight genes were expressed in the roots and these genes exhibited tissue- and cultivar-specific expression patterns.

Genome-wide identification of salinity responsive HSP70s in common bean.

The present study is aimed to identify and characterize HSP70 (PvHSP70) genes in two different common bean cultivars under salt stress. For this purpose various in silico methods such as RNAseq data and qRT-PCR analysis were used. A total of 24 candidate PvHSP70 gene were identified. Except for chromosome 4 and 7, these candidate PvHSP70 genes were distributed on the remaining chromosomes. While the lowest number of PvHSP70 genes was determined on chromosomes 1, 3, 5, 7, 9, 10 and 11 (one HSP70 gene), the highest number of PvHSP70s was on chromosomes 6 and 8 (seven HSP70 genes each). Three genes; PvHSP70-5, -9, and -10 were found to have no-introns. In addition, four tandemly and six segmentally duplicated gene couples were detected. A total of 13 PvHSP70 genes were targeted by miRNAs of 44 plant species and the most targeted genes were PvHSP70-5 and -23. The expression profile of PvHSP70 genes based on publicly available RNA-seq data was identified and salt treated leaf tissue was found to have more gene expression levels compared to the root. qRT-PCR analysis showed that the transcript concentrations of upregulated PvHSP70 genes in leaves of Zulbiye (sensitive) were mostly higher than those of Yakutiye (resistant). The present study revealed that PvHSP70 genes might play an important role in salt stress response for common bean cultivars and variability between cultivars also suggests that these genes could be used as functional markers for salt tolerance in common bean.

Roles of catalase (CAT) and ascorbate peroxidase (APX) genes in stress response of eggplant (Solanum melongena L.) against Cu(+2) and Zn(+2) heavy metal stresses.

Eggplant (Solanum melongena L.) is a good source of minerals and vitamins and this feature makes its value comparable with tomato which is economically the most important vegetable worldwide. Due to its common usage as food and in medicines, eggplant cultivation has a growing reputation worldwide. But genetic yield potential of an eggplant variety is not always attained, and it is limited by some factors such as heavy metal contaminated soils in today's world. Today, one of the main objectives of plant stress biology and agricultural biotechnology areas is to find the genes involved in antioxidant stress response and engineering the key genes to improve the plant resistance mechanisms. In this regard, the current study was conducted to gain an idea on the roles of catalase (CAT) and ascorbate peroxidase (APX) genes in defense mechanism of eggplant (S. melongena L., Pala-49 (Turkish cultivar)) treated with different concentrations of Cu(+2) and Zn(+2). For this aim, the steady-state messenger RNA (mRNA) levels of CAT and APX genes were determined by quantitative real-time PCR (qRT-PCR) in stressed eggplants. The results of the current study showed that different concentrations of Cu(+2) and Zn(+2) stresses altered the mRNA levels of CAT and APX genes in eggplants compared to the untreated control samples. When the mRNA levels of both genes were compared, it was observed that CAT gene was more active than APX gene in eggplant samples subjected to Cu(+2) contamination. The current study highlights the importance of CAT and APX genes in response to Cu(+2) and Zn(+2) heavy metal stresses in eggplant and gives an important knowledge about this complex interaction.

The determination of physiological and DNA changes in seedlings of maize (Zea mays L.) seeds exposed to the waters of the Gediz River and copper heavy metal stress.

In this study, the effects of the heavy metal-polluted waters of the Gediz River, which flow into the Aegean Sea, and different concentrations of copper (Cu) solutions on maize (Zea mays L.) seedlings are investigated with physiological parameters and random amplified polymorphic DNA (RAPD) assay. Results displayed physiologically a significant difference in root and stem length between the control seedlings and the seedlings grown with the waters of the Gediz River. Also, the certain ascending concentrations of copper solution (80, 160, 320, 640, and 1280 ppm) caused a significant decrease in root and stem length of seedlings compared to the control seedlings. As a result of the waters of the Gediz River and copper solution treatment, the changes occurred in RAPD profiles of seedlings observed as variations like increment and/or loss of bands compared with the control seedlings. And these changes were reflected as a decrease in genomic template stability (GTS, changes in RAPD profile) derived by genotoxicity. RAPD band profiles and GTS values showed consistent results with physiological parameter. In conclusion, the study revealed the environmental risk and negative effect of waters of the Gediz River on maize seedlings and the suitability of RAPD assay for the detection of environmental toxicology.

Characterization of stress induced by copper and zinc on cucumber (Cucumis sativus L.) seedlings by means of molecular and population parameters.

Contamination of plants with heavy metals could result in damage in DNA, such as mutations and cross-links with proteins. These altered DNA profiles may become visible in changes such as the appearance of a new band, or loss of an existing band, in the random amplified polymorphic DNA (RAPD) assay. In this study, various concentrations of copper and zinc salts were applied to cucumber seedlings during germination. Results displayed abnormalities in germination and also changes in root elongation, dry weight and total soluble protein level. All treatment concentrations (40, 80, 160, 240, 320, and 640mg/L) used in the study caused a decrease/delay in germination of the cucumbers to different extents. Inhibition or activation of root elongation was considered to be the first effect of metal toxicity in the tested plants. Application of the metal salts and the combined solutions on cucumber (Cucumis sativus L.) seedlings revealed similar consequences for total soluble protein level, dry weight and ultimately in inhibitory rates as well. The data obtained from RAPD band-profiles and genomic template stability (GTS) showed results that were consistent with the population parameters. In this regard, we conclude that molecular marker assays can be applied in combination with population parameters to measure genotoxic effects of heavy metals on plants.