Major Molecular Events Research Articles

Chemical Implications of apo-8, 6' Carotendial versus Intact Lycopene on Mechanism of Enhanced Cell-cell Communication and Apoptosis Induction in Breast Cancer Cells.

Investigation on carotenoids and its cleavage products is crucial to combat the development of chronic diseases, including cancer. Therefore, this study aimed to explore the effect of lycopene oxidative products versus equivalent concentration of lycopene (LYC) on major molecular events of cancer cells (MCF-7). Primarily, LYC-oxidized products were generated chemically, then collected its rich fraction. Based on cell-based assays, the antiproliferation potency of rich fraction of chemically-oxidized lycopene (COL) identified as apo-8, 6' carotendial was compared with LYC. Interestingly, the inhibition of cell migration by COL strongly demonstrated anti-metastatic activity. Further, the increased connexin-43 expression confirms enhanced gap-junctional communication activity of COL than LYC and control. Fortunately, apo-8, 6' carotendial did not affect normal breast epithelial cells. We anticipated that, the chemical properties of apo-8, 6'-carotendial is similar and mimic a model compound acrolein (α, β-conjugated aldehyde) which is involved in Michael addition/Schiff base formation with specific amino acids and regulates redox signaling, reactive oxygen species sensing and cellular buffering. The chemistry of apo-8, 6' carotendial reveals a greater insight into the mechanism of selective inhibition of cancer cells proliferation. In this context, speculations of putative action of lycopeneoids through chemical biology approach facilitate greater insights in tandem with synthetic chemistry.

Assessment of Photosensitizer Concentration Effects on the Efficacy of Photodynamic Therapy.

Introduction: Photodynamic therapy (PDT) is an attractive approach in medicine. Due to its noninvasive nature and low side effects, PDT has been developed quickly. In the present study, the gene expression profiles of the human cell line that was treated via PDT in the sub-lethal concentration (LC50) and super-lethal concentration (LC90) of a photosensitizer (PS) from Gene Expression Omnibus (GEO) were extracted and the common differentially expressed genes (DEGs) were investigated. Methods: The gene expression profiles of the treated cells were compared with a control, and the common DEGs were determined. The common DEGs were assessed via protein-protein interaction (PPI) network analysis, and gene ontology enrichment was evaluated. The related biological terms for the common genes were identified. Results: Ninety-four common DEGs were selected to be analyzed. It appeared that the activation and increment of gene expression were prominent processes. Jun, Dusp1, Atf4, and Atf3 as four critical genes were highlighted. "Chromosomal and microsatellite instability in colorectal cancer" was identified as the main class of biological terms related to the assessed DEGs. Conclusion: The major molecular events which happened in both analyses indicated that PDT, independent from the concentration of PS, induced gross molecular changes such as the upregulation of Jun and Dusp1.

Cardiac Hypoxia Tolerance in Fish: From Functional Responses to Cell Signals.

Aquatic animals are increasingly challenged by O2 fluctuations as a result of global warming, as well as eutrophication processes. Teleost fish show important species-specific adaptability to O2 deprivation, moving from intolerance to a full tolerance of hypoxia and even anoxia. An example is provided by members of Cyprinidae which includes species that are amongst the most tolerant hypoxia/anoxia teleosts. Living at low water O2 requires the mandatory preservation of the cardiac function to support the metabolic and hemodynamic requirements of organ and tissues which sustain whole organism performance. A number of orchestrated events, from metabolism to behavior, converge to shape the heart response to the restricted availability of the gas, also limiting the potential damages for cells and tissues. In cyprinids, the heart is extraordinarily able to activate peculiar strategies of functional preservation. Accordingly, by using these teleosts as models of tolerance to low O2, we will synthesize and discuss literature data to describe the functional changes, and the major molecular events that allow the heart of these fish to sustain adaptability to O2 deprivation. By crossing the boundaries of basic research and environmental physiology, this information may be of interest also in a translational perspective, and in the context of conservative physiology, in which the output of the research is applicable to environmental management and decision making.

Chromosome 20q and 13q gain in metastatic colorectal cancer: Prognostic significance and genomic correlates.

e15553 Background: Chromosome 13q and 20q gain are major molecular events in adenoma-carcinoma progression and metastasis. The prognostic significance and correlates of 13q and 20q gain in metastatic colorectal cancer (CRC) have not been established. Methods: A total of 108 cases were selected at the University of Louisville. We ran a custom research-use only algorithm to assess whole arm chr13q and chr20q gain from next generation sequencing data of CRC samples. The algorithm utilized the coverage data of baited regions of the genome to model the copy number of each segment. A gain call was made if > 50% of the chromosome arm was gained. Tumors were noted as left-sided colon, right-sided colon, or rectal cancers. Data on demographics, treatment and Kaplan-Meier survival were analyzed. Results: The prevalence of 13q gain was 58% and 20q gain was detected in 61% of cases. Concurrent 13q and 20q gains were frequent, with 49% prevalence. The only 2 MSI-High samples were negative for 13q or 20q gain. Tumors with 13q and 20q gain were significantly more likely to have mutations in TP53 and APC (Table). Rectal tumors were significantly enriched for 13q gain ( p= 0.020) and 20q gain ( p= 0.007). Examining survival by tumor location, 13q gain conferred significantly worse outcome for rectal cancers ( p= 0.007), as did 20q gain ( p= 0.037). Conclusions: With median follow up of over 3 years, 13q and 20q gains were significantly associated with worse prognosis in rectal cancers. The data indicate that 13q/20q gain and associated molecular signature are important prognostic markers in CRC.[Table: see text]

The SZT2 Interactome Unravels New Functions of the KICSTOR Complex.

Seizure threshold 2 (SZT2) is a component of the KICSTOR complex which, under catabolic conditions, functions as a negative regulator in the amino acid-sensing branch of mTORC1. Mutations in this gene cause a severe neurodevelopmental and epileptic encephalopathy whose main symptoms include epilepsy, intellectual disability, and macrocephaly. As SZT2 remains one of the least characterized regulators of mTORC1, in this work we performed a systematic interactome analysis under catabolic and anabolic conditions. Besides numerous mTORC1 and AMPK signaling components, we identified clusters of proteins related to autophagy, ciliogenesis regulation, neurogenesis, and neurodegenerative processes. Moreover, analysis of SZT2 ablated cells revealed increased mTORC1 signaling activation that could be reversed by Rapamycin or Torin treatments. Strikingly, SZT2 KO cells also exhibited higher levels of autophagic components, independent of the physiological conditions tested. These results are consistent with our interactome data, in which we detected an enriched pool of selective autophagy receptors/regulators. Moreover, preliminary analyses indicated that SZT2 alters ciliogenesis. Overall, the data presented form the basis to comprehensively investigate the physiological functions of SZT2 that could explain major molecular events in the pathophysiology of developmental and epileptic encephalopathy in patients with SZT2 mutations.

Single Cell RNA-Sequencing Reveals a Murine Gallbladder Cell Transcriptome Atlas During the Process of Cholesterol Gallstone Formation.

Gallstone disease is a worldwide common disease. However, the knowledge concerning the gallbladder in the pathogenesis of cholesterol gallstone formation remains limited. In this study, using single-cell RNA sequencing (scRNA-seq) to obtain the transcriptome of gallbladder cells, we showed cellular heterogeneity and transcriptomic dynamics in murine gallbladder cells during the process of lithogenesis. Our results indicated gallbladder walls were subjected to remodeling during the process of lithogenesis. The major molecular events that happened included proliferation of epithelial cells, infiltration of immune-cells, activation of angiogenesis, and extracellular matrix modulation. Furthermore, we observed partial reversal of gallbladder cell transcriptomes by ursodeoxycholic acid treatment. This work thus provides novel and integral knowledges on the cellular changes during lithogenesis, which is of great significance to the understanding of pathogenesis and treatment of cholesterol gallstone.

PP-1β and PP-2Aα modulate cAMP response element-binding protein (CREB) functions in aging control and stress response through de-regulation of αB-crystallin gene and p300-p53 signaling axis.

The function of the transcription factor, cAMP response element‐binding protein (CREB), is activated through S133 phosphorylation by PKA and others. Regarding its inactivation, it is not well defined. cAMP response element‐binding protein plays an essential role in promoting cell proliferation, neuronal survival and the synaptic plasticity associated with long‐term memory. Our recent studies have shown that CREB is an important player in mediating stress response. Here, we have demonstrated that CREB regulates aging process through suppression of αB‐crystallin and activation of the p300‐p53‐Bak/Bax signaling axis. First, we determined that two specific protein phosphatases, PP‐1β and PP‐2Aα, can inactivate CREB through S133 dephosphorylation. Subsequently, we demonstrated that cells expressing the S133A‐CREB, a mutant mimicking constant dephosphorylation at S133, suppress CREB functions in aging control and stress response. Mechanistically, S133A‐CREB not only significantly suppresses CREB control of αB‐crystallin gene, but also represses CREB‐mediated activation of p53 acetylation and downstream Bak/Bax genes. cAMP response element‐binding protein suppression of αB‐crystallin and its activation of p53 acetylation are major molecular events observed in human cataractous lenses of different age groups. Together, our results demonstrate that PP‐1β and PP‐2Aα modulate CREB functions in aging control and stress response through de‐regulation of αB‐crystallin gene and p300‐p53‐Bax/Bak signaling axis, which regulates human cataractogenesis in the aging lens.

Tribbles Homolog 3 Mediates the Development and Progression of Diabetic Retinopathy.

The current understanding of the molecular pathogenesis of diabetic retinopathy does not provide a mechanistic link between early molecular changes and the subsequent progression of the disease. In this study, we found that human diabetic retinas overexpressed TRIB3 and investigated the role of TRIB3 in diabetic retinal pathobiology in mice. We discovered that TRIB3 controlled major molecular events in early diabetic retinas via HIF1α-mediated regulation of retinal glucose flux, reprogramming cellular metabolism, and governing of inflammatory gene expression. These early molecular events further defined the development of neurovascular deficit observed in mice with diabetic retinopathy. TRIB3 ablation in the streptozotocin-induced mouse model led to significant retinal ganglion cell survival and functional restoration accompanied by a dramatic reduction in pericyte loss and acellular capillary formation. Under hypoxic conditions, TRIB3 contributed to advanced proliferative stages by significant upregulation of GFAP and VEGF expression, thus controlling gliosis and aberrant vascularization in oxygen-induced retinopathy mouse retinas. Overall, our data reveal that TRIB3 is a master regulator of diabetic retinal pathophysiology that may accelerate the onset and progression of diabetic retinopathy to proliferative stages in humans and present TRIB3 as a potentially novel therapeutic target for diabetic retinopathy.

Glutathione S-transferase gene polymorphic sequence variations: Association with risk and response to Imatinib among Chronic Myeloid Leukemia patients of Kashmir.

Glutathione S-transferase (GST) gene deletion or polymorphic sequence variations lead to decreased enzyme activity that influences susceptibility and response to tyrosine kinase inhibitors in chronic myeloid leukemia (CML). We aimed to analyze relation of different GST gene sequence variants with susceptibility and response to Imatinib in CML. A total of 150CML cases and equal number of age and gender matched healthy controls were genotyped for five GST polymorphisms by multiplex-PCR and PCR-RFLP techniques. BCR-ABL1 transcripts were quantified by quantitative Real Time PCR (qRT-PCR). GSTT1, GSTO1, and GSTO2 SNPs revealed no association, while as GSTM1null genotype was observed to protect against the development of CML (OR=0.53, P=.01). GSTP1 variant genotypes AG (OR=2.1, P=.003) and GG (OR=5.6, P<.001), significantly associated with increased risk of CML. Combined genotype analysis showed protective impact of GSTT1present /GSTM1null (OR=0.44, P=.003) while as GSTT1present /GSTP1-GG (OR=6.92, P<.001) and GSTM1present /GSTP1-GG (OR=6.33, P<.001), significantly increased CML risk. GSTM1null genotype individually and in combination with GSTT1present associated with superior rate of major molecular response (MMR) and event free survival (EFS) (log-rank P=.029). GSTO2-AG+GG genotype associated with significantly inferior MMR rates at 3, 6, and 12months. Also, patients with GSTO2-GG genotype showed significantly reduced EFS (log-rank P=.025). Multivariate analysis confirmed GSTM1null as a better (HR:0.19, P=.029) and GSTO2-GG genotype as an independent poor prognostic factor (HR:2.29, P=.037). GSTM1null genotype seems to have a better prognostic role while GSTP1 variants significantly increase CML risk. Also, results support a correlation between disease outcome and GSTO2 polymorphism.

Dissection of the Ovulatory Process Using ex vivo Approaches.

Ovulation is a unique physiological phenomenon that is essential for sexual reproduction. It refers to the entire process of ovarian follicle responses to hormonal stimulation resulting in the release of mature fertilization-competent oocytes from the follicles and ovaries. Remarkably, ovulation in different species can be reproduced out-of-body with high fidelity. Moreover, most of the molecular mechanisms and signaling pathways engaged in this process have been delineated using in vitro ovulation models. Here, we provide an overview of the major molecular and cytological events of ovulation observed in frogs, primarily in the African clawed frog Xenopus laevis, using mainly ex vivo approaches, with the focus on meiotic oocyte maturation and follicle rupture. For the purpose of comparison and generalization, we also refer extensively to ovulation in other biological species, most notoriously, in mammals.

Protein Kinase C inhibitor Molecule Effect Partial Reversal of Spliceopathy in Myotonic Dystrophy Type 2 Cell Line

Myotonic dystrophy is a neuromuscular disease which manifest as two forms namely type 1 (DM1) and type 2 (DM2). One of the major molecular events associated with the disease condition is misregulation of splicing resulting in spliceopathy. This study was performed to assess the effect of protein kinase C (PKC) inhibitors- Ro 31-8220 and hypericin- on the splicing of insulin receptor (IR) and muscleblind-like (MBNL1) in myotonic dystrophy type 2 (DM2) cell line. The results of this study demonstrated that only Ro 31-8820 was able to effect partial rescue of missplicing of insulin receptor and muscleblind-like 1. It indicates that it could be a possible therapeutic agent for treatment of DM2. Keywords : Myotonic dystrophy, Protein Kinase C inhibitor, spliceopathy, therapeutics DOI: 10.7176/JNSR/11-14-03 Publication date: July 31 st 2020

Localization shift of a sugar transporter contributes to phloem unloading in sweet watermelons.

Unloading sugar from sink phloem by transporters is complex and much remains to be understood about this phenomenon in the watermelon fruit. Here, we report a novel vacuolar sugar transporter (ClVST1) identified through map-based cloning and association study, whose expression in fruit phloem is associated with accumulation of sucrose (Suc) in watermelon fruit. ClVST197 knockout lines show decreased sugar content and total biomass, whereas overexpression of ClVST197 increases Suc content. Population genomic and subcellular localization analyses strongly suggest a single-base change at the coding region of ClVST197 as a major molecular event during watermelon domestication, which results in the truncation of 45 amino acids and shifts the localization of ClVST197 to plasma membranes in sweet watermelons. Molecular, biochemical and phenotypic analyses indicate that ClVST197 is a novel sugar transporter for Suc and glucose efflux and unloading. Functional characterization of ClVST1 provides a novel strategy to increase sugar sink potency during watermelon domestication.

Quantitative mass spectrometric analysis of the mouse cerebral cortex after ischemic stroke.

Ischemic strokes result in the death of brain tissue and a wave of downstream effects, often leading to lifelong disabilities or death. However, the underlying mechanisms of ischemic damage and repair systems remain largely unknown. In order to better understand these mechanisms, TMT-isobaric mass tagging and mass spectrometry were conducted on brain cortex extracts from mice subjected to one hour of middle cerebral artery occlusion (MCAO) and after one hour of reperfusion. In total, 2,690 proteins were identified and quantified, out of which 65% of the top 5% of up- and down-regulated proteins were found to be significant (p < 0.05). Network-based gene ontology analysis was then utilized to cluster all identified proteins by protein functional groups and cellular roles. Although three different cellular functions were identified—organelle outer membrane proteins, cytosolic ribosome proteins, and spliceosome complex proteins—several functional domains were found to be common. Of these, organelle outer membrane proteins were downregulated whereas cytosolic ribosome and spliceosome complex proteins were upregulated, indicating that major molecular events post-stroke were translation-associated and subsequent signaling pathways (e.g., poly (ADP-ribose) (PAR) dependent cell death). By approaching stroke analyses via TMT-isobaric mass tagging, the work herein presents a grand scope of protein-based molecular mechanisms involved with ischemic stroke recovery.

Tau hyperphosphorylation induced by the anesthetic agent ketamine/xylazine involved the calmodulin-dependent protein kinase II.

Tau hyperphosphorylation is a major neuropathological hallmark of many neurodegenerative disorders such as Alzheimer's disease. Several anesthetics have been shown previously to induced marked tau hyperphosphorylation. Although the ketamine/xylazine mixture is one of the most commonly used anesthetic agents in animal research and veterinary practice, the effect of this anesthetic agent on tau phosphorylation still remains to be determined. Here, we found that ketamine-/xylazine-induced a rapid and robust hyperphosphorylation of tau in a dose-dependent manner under normothermic and hypothermic conditions in mice. When used together, ketamine and xylazine exerted a synergistic action on tau phosphorylation most strongly not only on epitopes S396 and S262, but also on other residues (T181, and S202/T205). We observed that activation of the calmodulin-dependent protein kinase II (CaMKII) is the major upstream molecular event leading to tau hyperphosphorylation following ketamine/xylazine anesthesia in mice. Moreover, we observed that intracerebroventricular injection of the selective CaMKII inhibitor KN93 attenuated tau hyperphosphorylation. Since ketamine/xylazine also had a marked impact on other key molecular signaling pathways involving the MAP/microtubule affinity-regulating kinase (MARK), extracellular signal-regulated kinase (ERK), and glycogen synthase kinase-3 (GSK3), our study calls for high caution and careful monitoring when using this anesthetic agent in laboratory animal settings across all fields of biological sciences in order to avoid artifactual results.

Inherent variations in the cellular events at the site of amputation orchestrate scar-free wound healing in the tail and scarred wound healing in the limb of lizard Hemidactylus flaviviridis.

Lizards are unique in having both-regeneration competent (tail) as well as non-regenerating appendages (limbs) in adults. They therefore present an appropriate model for comparing processes underlying regenerative repair and nonregenerative healing after amputation. In the current study, we use northern house gecko Hemidactylus flaviviridis to compare major cellular and molecular events following amputation of the limb and of the tail. Although the early response to injury in both cases comprises apoptosis, proliferation, and angiogenesis, the temporal distribution of these processes in each remained obscure. In this regard, observations were made on the anatomy and gene expression levels of key regulators of these processes during the healing phase of the tail and limb separately. It was revealed that cell proliferation markers like fibroblast growth factors were upregulated early in the healing tail, coinciding with the growing epithelium. The amputated limb, in contrast, showed weak expression of proliferation markers, limited only to fibroblasts in the later stage of healing. Additionally, apoptotic activity in the tail was limited to the very early phase of healing, as opposed to that in the limb, wherein high expression of caspase-3 was observed throughout the healing process. Early rise in VEGF-α expression reflected an early onset of angiogenesis in the tail, while it was seen to occur at a later stage in case of the limb. Moreover, the expression pattern of transforming growth factor beta members points toward a pro-fibrotic response being induced very early in the amputated limb. Collectively, these results explain why regenerating appendages are able to heal without scars and if we are to induce scar-free healing in nonregenerating limbs, what interventions can be envisaged. This is crucial to the field of regenerative medicine since it is the initial stages of repair following amputation, which decide whether the appendage will be restored or only covered with a scab.

Combination therapy with KRAS siRNA and EGFR inhibitor AZD8931 suppresses lung cancer cell growth in vitro.

Lung cancer is a leading cause of cancer-related deaths worldwide, with less than a 5-year survival rate for both men and women. Epidermal growth factor receptor (EGFR) and Kirsten rat sarcoma oncogene (KRAS) signaling pathways play a critical role in the proliferation and progression of various cancers, including lung cancer. Genetic studies have shown that amplification, over-expression, or mutation of EGFR is an early and major molecular event in many human tumors. KRAS mutation is a negative factor in various cancer, including non-small-cell lung cancer, and complicates therapeutic approaches with adjuvant chemotherapy and anti-EGFR directed therapies. This article is dedicated to evaluating the synergistic effect of a novel EGFR inhibitor AZD8931 and KRAS small interfering RNA (siRNA) on the proliferation and apoptosis of lung adenocarcinoma cancer cells. A549 lung cancer cells were treated with KRAS siRNA and the EGFR inhibitor alone or in combination. The cytotoxic effects of KRAS siRNA and te EGFR inhibitor were determined usingMTT assay, and induction of apoptosis was determined by FACS analysis. Suppression of KRAS, Her-2, and EGFR expression by treatments was measured by qRT-PCR and western blotting.KRAS siRNA and the EGFR inhibitor significantly reduced the proliferation of A549 cells as well as KRAS and EGFR mRNA levels 24 hr after treatment. The results also indicated that the silencing of KRAS and EGFR has synergistic effects on the induction of apoptosis on the A549 cells.These results indicated that KRAS and EGFR might play important roles in the progression of lung cancer and could be potential therapeutic targets for treatment of lung cancer.

U1 snRNP Alteration and Neuronal Cell Cycle Reentry in Alzheimer Disease.

The aberrancy of U1 small nuclear ribonucleoprotein (snRNP) complex and RNA splicing has been demonstrated in Alzheimer’s disease (AD). Importantly, the U1 proteopathy is AD-specific, widespread and early-occurring, thus providing a very unique clue to the AD pathogenesis. The prominent feature of U1 histopathology is its nuclear depletion and redistribution in the neuronal cytoplasm. According to the preliminary data, the initial U1 cytoplasmic distribution pattern is similar to the subcellular translocation of the spliceosome in cells undergoing mitosis. This implies that the U1 mislocalization might reflect the neuronal cell cycle-reentry (CCR) which has been extensively evidenced in AD brains. The CCR phenomenon explains the major molecular and cellular events in AD brains, such as Tau and amyloid precursor protein (APP) phosphorylation, and the possible neuronal death through mitotic catastrophe (MC). Furthermore, the CCR might be mechanistically linked to inflammation, a critical factor in the AD etiology according to the genetic evidence. Therefore, the discovery of U1 aberrancy might strengthen the involvement of CCR in the AD neuronal degeneration.

Acquisition of oocyte competence to develop as an embryo: integrated nuclear and cytoplasmic events.

Infertility affects ~7% of couples of reproductive age with little change in incidence in the last two decades. ART, as well as other interventions, have made major strides in correcting this condition. However, and in spite of advancements in the field, the age of the female partner remains a main factor for a successful outcome. A better understanding of the final stages of gamete maturation yielding an egg that can sustain embryo development and a pregnancy to term remains a major area for improvement in the field. This review will summarize the major cellular and molecular events unfolding at the oocyte-to-embryo transition. We will provide an update on the most important processes/pathways currently understood as the basis of developmental competence, including the molecular processes involved in mRNA storage, its recruitment to the translational machinery, and its degradation. We will discuss the hypothesis that the translational programme of maternal mRNAs plays a key role in establishing developmental competence. These regulations are essential to assemble the machinery that is used to establish a totipotent zygote. This hypothesis further supports the view that embryogenesis begins during oogenesis. A better understanding of the events required for developmental competence will guide the development of novel strategies to monitor and improve the success rate of IVF. Using this information, it will be possible to develop new biomarkers that may be used to better predict oocyte quality and in selection of the best egg for IVF.

The Effect of Beta-Amyloid Peptides and Main Stress Protein HSP70 on Human SH-SY5Y Neuroblastoma Proteome

The accumulation and aggregation of β-amyloids are major molecular events underlying the progression of Alzheimer's disease. In neural cells, recombinant HSP70 reduces the toxic effect of Aβ and its isomeric forms. Here we describe the proteome of the neuroblastoma cell line after incubation with amyloid peptides Aβ42 and isomerized Asp7 (isoAβ42) without and with human recombinant heat shock protein 70 (HSP70). Incubation of SH-SY5Y cell culture with the synthetic Aβ-peptides leads to a decrease in the levels of several cytoskeleton proteins (e.g., ACTN1, VIME, TPM3) and several chaperonines involved in the folding of actin and tubulin (TCPQ, TCPG, TCPE, TCPB). These changes are accompanied by an increase in the expression of calmodulin and the proteins involved in folding in the endoplasmic reticulum and endoplasmic cell stress response. The presence of exogenous HSP70 has led to an increase in expression of several chaperones and a few other proteins including endogenous HSP70. A combined effect of recombinant HSP70 with Aβ peptides reduced cell apoptosis and significantly decreased the level of tubulin phosphorylation caused by the addition of Aβ peptides.

A Tonoplast Sugar Transporter Underlies a Sugar Accumulation QTL in Watermelon.

How sugar transporters regulate sugar accumulation in fruits is poorly understood and particularly so for species storing high-concentration Suc. Accumulation of soluble sugars in watermelon (Citrullus lanatus) fruit, a major quality trait, had been selected during domestication. Still, the molecular mechanisms controlling this quantitative trait are unknown. We resequenced 96 recombinant inbred lines, derived from crossing sweet and unsweet accessions, to narrow down the size of a previously described sugar content quantitative trait locus, which contains a putative Tonoplast Sugar Transporter gene (ClTST2). Molecular and biochemical analyses indicated that ClTST2 encodes a vacuolar membrane protein, whose expression is associated with tonoplast uptake and accumulation of sugars in watermelon fruit flesh cells. We measured fruit sugar content and resequenced the genomic region surrounding ClTST2 in 400 watermelon accessions and associated the most sugar-related significant single-nucleotide polymorphisms (SNPs) to the ClTST2 promoter. Large-scale population analyses strongly suggest increased expression of ClTST2 as a major molecular event in watermelon domestication associated with a selection sweep around the ClTST2 promoter. Further molecular analyses explored the binding of a sugar-induced transcription factor (SUSIWM1) to a sugar-responsive cis-element within the ClTST2 promoter, which contains the quantitative trait locus (QTL) causal SNP. The functional characterization of ClTST2 and its expression regulation by SUSIWM1 provide novel tools to increase sugar sink potency in watermelon and possibly in other vegetable and fruit crops.