Search Results (12)

Melanoma is one of the most aggressive and fatal cancers; however, effective and long-lasting treatment options for melanoma continue to be sought after due to the development of resistance mechanisms to the currently available therapies. Background: The K-homology-type splicing regulatory protein (KSRP) is an RNA-binding regulatory protein that binds to the AU-rich elements at the 3′-UTR of target mRNAs. Prior studies have demonstrated that KSRP plays a crucial role in the post-transcriptional regulation of gene expression in human melanoma. Subsequently, in this study, we further examined the role of KSRP in cell migration, colony formation, apoptosis, and tumorigenicity of human melanoma. Methods: KSRP was knocked down in two different human melanoma cell lines: A375 and SK-MEL-28, using lenti-shRNA techniques. By doing so, we studied the effects of KSRP inhibition on cell migration, colony formation, proliferation, apoptosis, and tumorigenicity in these melanoma cell lines. Results: We observed a significant decrease in cell migration, colony formation, proliferation, and tumorigenicity, while also observing a substantial increase in apoptosis in the KSRP knock down melanoma cell lines. Conclusions: Our data establishes that KSRP plays a vital role in cell migration, colony formation, proliferation, apoptosis, and tumorigenicity in both the A375 and SK-MEL-28 human melanoma cell lines. Full article

The mRNA-binding protein KSRP (KH-type splicing regulatory protein) is known to modulate immune cell functions post-transcriptionally, e.g., by reducing the mRNA stability of cytokines. It is known that KSRP binds the AU-rich motifs (ARE) that are often located in the 3′-untranslated part of mRNA species, encoding dynamically regulated proteins as, for example, cytokines. Innate myeloid immune cells, such as polymorphonuclear neutrophils (PMNs) and macrophages (MACs), eliminate pathogens by multiple mechanisms, including phagocytosis and the secretion of chemo- and cytokines. Here, we investigated the role of KSRP in the phenotype and functions of both innate immune cell types in the mouse model of invasive pulmonary aspergillosis (IPA). Here, KSRP^−/− mice showed lower levels of Aspergillus fumigatus conidia (AFC) and an increase in the frequencies of PMNs and MACs in the lungs. Our results showed that PMNs and MACs from KSRP^−/− mice exhibited an enhanced phagocytic uptake of AFC, accompanied by increased ROS production in PMNs upon stimulation. A comparison of RNA sequencing data revealed that 64 genes related to inflammatory and immune responses were shared between PMNs and MACs. The majority of genes upregulated in PMNs were involved in metabolic processes, cell cycles, and DNA repair. Similarly, KSRP-deficient PMNs displayed reduced levels of apoptosis. In conclusion, our results indicate that KSRP serves as a critical negative regulator of PMN and MAC anti-pathogen activity. Full article

Transcriptional regulation constitutes a key step in gene expression regulation. Myocyte enhancer factor 2C (MEF2C) is a transcription factor of the MADS box family involved in the early development of several cell types, including muscle cells. Over the last decade, a novel layer of complexity modulating gene regulation has emerged as non-coding RNAs have been identified, impacting both transcriptional and post-transcriptional regulation. microRNAs represent the most studied and abundantly expressed subtype of small non-coding RNAs, and their functional roles have been widely documented. On the other hand, our knowledge of the transcriptional and post-transcriptional regulatory mechanisms that drive microRNA expression is still incipient. We recently demonstrated that MEF2C is able to transactivate the long, but not short, regulatory element upstream of the miR-23a-miR-27a-miR-24-2 transcriptional start site. However, MEF2C over-expression and silencing, respectively, displayed distinct effects on each of the miR-23a-miR-27a-miR-24-2 mature cluster members without affecting pri-miRNA expression levels, thus supporting additional MEF2C-driven regulatory mechanisms. Within this study, we demonstrated a complex post-transcriptional regulatory mechanism directed by MEF2C in the regulation of miR-23a-miR-27a-miR-24-2 cluster members, distinctly involving different domains of the MEF2C transcription factor and the physical interaction with pre-miRNAs and Ksrp, HnRNPa3 and Ddx17 transcripts. Full article

KH-type splicing regulatory protein (KSRP) is a single-stranded nucleic acid-binding protein with multiple functions. It is known to bind AU-rich motifs within the 3′-untranslated region of mRNA species, which in many cases encode dynamically regulated proteins like cytokines. In the present study, we investigated the role of KSRP for the immunophenotype of macrophages using bone marrow-derived macrophages (BMDM) from wild-type (WT) and KSRP^−/− mice. RNA sequencing revealed that KSRP^−/− BMDM displayed significantly higher mRNA expression levels of genes involved in inflammatory and immune responses, particularly type I interferon responses, following LPS stimulation. In line, time kinetics studies revealed increased levels of interferon-γ (IFN-γ), interleukin (IL)-1β and IL-6 mRNA in KSRP^−/− macrophages after 6 h subsequent to LPS stimulation as compared to WT cultures. At the protein level, KSRP^−/− BMDM displayed higher levels of these cytokines after overnight stimulation. Matching results were observed for primary peritoneal macrophages of KSRP^−/− mice. These showed higher IL-6, tumor necrosis factor-α (TNF-α), C-X-C motif chemokine 1 (CXCL1) and CC-chemokine ligand 5 (CCL5) protein levels in response to LPS stimulation than the WT controls. As macrophages play a key role in sepsis, the in vivo relevance of KSRP deficiency for cytokine/chemokine production was analyzed in an acute inflammation model. In agreement with our in vitro findings, KSRP-deficient animals showed higher cytokine production upon LPS administration in comparison to WT mice. Taken together, these findings demonstrate that KSRP constitutes an important negative regulator of cytokine expression in macrophages. Full article

The obesity epidemic represents a critical public health issue worldwide, as it is a vital risk factor for many diseases, including type 2 diabetes (T2D) and cardiovascular disease. Obesity is a complex disease involving excessive fat accumulation. Proper adipose tissue accumulation and function are highly transcriptional and regulated by many genes. Recent studies have discovered that post-transcriptional regulation, mainly mediated by RNA-binding proteins (RBPs), also plays a crucial role. In the lifetime of RNA, it is bound by various RBPs that determine every step of RNA metabolism, from RNA processing to alternative splicing, nucleus export, rate of translation, and finally decay. In humans, it is predicted that RBPs account for more than 10% of proteins based on the presence of RNA-binding domains. However, only very few RBPs have been studied in adipose tissue. The primary aim of this paper is to provide an overview of RBPs in adipogenesis and adipose function. Specifically, the following best-characterized RBPs will be discussed, including HuR, PSPC1, Sam68, RBM4, Ybx1, Ybx2, IGF2BP2, and KSRP. Characterization of these proteins will increase our understanding of the regulatory mechanisms of RBPs in adipogenesis and provide clues for the etiology and pathology of adipose-tissue-related diseases. Full article

Karst aquifers are both a valuable resource for humankind and a habitat for unique biota. The quality of freshwater sources may be easily affected by natural (e.g., geology, climate, and vegetation) and anthropogenic (e.g., agriculture, livestock, and tourism) changes, particularly in karst landscapes with highly vulnerable groundwater reservoirs. We seasonally monitored nine representative freshwater sources (i.e., six springs, a well, a surface stream, and a cave stream resurgence) in the karst system of the Runcuri Plateau (KSRP) (Western Romanian Carpathians) during seven sampling campaigns in 2019–2021. We assessed how these natural and anthropogenic factors influenced the water quality based on the European and national standards for drinking water. The geological structure (i.e., tectonics and lithology) of the KSRP was reassessed, and the environmental variables of the freshwater sites were investigated in order to evaluate their impact on the physicochemical profile, the microbial contamination, and on the meiofauna presence. Multivariate statistics were performed to gain insights into the interplay among all these factors and to evaluate the self-purification capacity of the KSRP for chemical and microbial pollutants. The most relevant drivers shaping the microbial content of the freshwater sources were the altitude of the sampling sites, the normalized difference vegetation index (NDVI), and air temperature, followed by the physicochemical profile of the waters (i.e., calcium hardness, magnesium hardness, nitrites, nitrates, conductivity, phosphates, total dissolved solids, and iron concentrations). The meiofauna presence was influenced mostly by precipitation, air temperature, and NDVI. Our results reflected the effect of the geological structure and environment on water chemistry and biota assemblages. A pollutant attenuation trend was observed in discharging waters, even though the self-purification capacity of the studied karst system was not statistically supported. More investigations are needed to comprehend the processes developed in the black box of the KSRP. Full article

Post-transcriptional control of gene expression is one important mechanism that enables stringent and rapid modulation of cytokine, chemokines or growth factors expression, all relevant for immune or tumor cell function and communication. The RNA-binding protein KH-type splicing regulatory protein (KSRP) controls the mRNA stability of according genes by initiation of mRNA decay and inhibition of translation, and by enhancing the maturation of microRNAs. Therefore, KSRP plays a pivotal role in immune cell function and tumor progression. In this review, we summarize the current knowledge about KSRP with regard to the regulation of immunologically relevant targets, and the functional role of KSRP on immune responses and tumorigenesis. KSRP is involved in the control of myeloid hematopoiesis. Further, KSRP-mediated mRNA decay of pro-inflammatory factors is necessary to keep immune homeostasis. In case of infection, functional impairment of KSRP is important for the induction of robust immune responses. In this regard, KSRP seems to primarily dampen T helper cell 2 immune responses. In cancer, KSRP has often been associated with tumor growth and metastasis. In summary, aside of initiation of mRNA decay, the KSRP-mediated regulation of microRNA maturation seems to be especially important for its diverse biological functions, which warrants further in-depth examination. Full article

Secondary hyperparathyroidism (SHP) is a common complication of chronic kidney disease (CKD) that induces morbidity and mortality in patients. How CKD stimulates the parathyroid to increase parathyroid hormone (PTH) secretion, gene expression and cell proliferation remains an open question. In experimental SHP, the increased PTH gene expression is post-transcriptional and mediated by PTH mRNA–protein interactions that promote PTH mRNA stability. These interactions are orchestrated by the isomerase Pin1. Pin1 participates in conformational change-based regulation of target proteins, including mRNA-binding proteins. In SHP, Pin1 isomerase activity is decreased, and thus, the Pin1 target and PTH mRNA destabilizing protein KSRP fails to bind PTH mRNA, increasing PTH mRNA stability and levels. An additional level of post-transcriptional regulation is mediated by microRNA (miRNA). Mice with parathyroid-specific knockout of Dicer, which facilitates the final step in miRNA maturation, lack parathyroid miRNAs but have normal PTH and calcium levels. Surprisingly, these mice fail to increase serum PTH in response to hypocalcemia or uremia, indicating a role for miRNAs in parathyroid stimulation. SHP often leads to parathyroid hyperplasia. Reduced expressions of parathyroid regulating receptors, activation of transforming growth factor α-epidermal growth factor receptor, cyclooxygenase 2-prostaglandin E2 and mTOR signaling all contribute to the enhanced parathyroid cell proliferation. Inhibition of mTOR by rapamycin prevents and corrects the increased parathyroid cell proliferation of SHP. This review summarizes the current knowledge on the mechanisms that stimulate the parathyroid cell at multiple levels in SHP. Full article

KH-type splicing regulatory protein (KSRP) is an RNA-binding protein that promotes mRNA decay and thereby negatively regulates cytokine expression at the post-transcriptional level. Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by dysregulated cytokine expression causing multiple organ manifestations; MRL-Fas^lpr mice are an established mouse model to study lupus disease pathogenesis. To investigate the impact of KSRP on lupus disease progression, we generated KSRP-deficient MRL-Fas^lpr mice (MRL-Fas^lpr/KSRP^−/− mice). In line with the predicted role of KSRP as a negative regulator of cytokine expression, lupus nephritis was augmented in MRL-Fas^lpr/KSRP^−/− mice. Increased infiltration of immune cells, especially of IFN-γ producing T cells and macrophages, driven by enhanced expression of T cell-attracting chemokines and adhesion molecules, seems to be responsible for worsened kidney morphology. Reduced expression of the anti-inflammatory interleukin-1 receptor antagonist may be another reason for severe inflammation. The increase of FoxP3⁺ T cells detected in the kidney seems unable to dampen the massive kidney inflammation. Interestingly, lymphadenopathy was reduced in MRL-Fas^lpr/KSRP^−/− mice. Altogether, KSRP appears to have a complex role in immune regulation; however, it is clearly able to ameliorate lupus nephritis. Full article

Transcripts of alpha-fetoprotein (Afp), H19, and insulin-like growth factor 2 (Igf2) genes are highly expressed in mouse fetal liver, but decrease drastically during maturation. While transcriptional regulation of these genes has been well studied, the post-transcriptional regulation of their developmental decrease is poorly understood. Here, we show that shortening of poly(A) tails and subsequent RNA decay are largely responsible for the postnatal decrease of Afp, H19, and Igf2 transcripts in mouse liver. IGF2 mRNA binding protein 1 (IMP1), which regulates stability and translation efficiency of target mRNAs, binds to these fetal liver transcripts. When IMP1 is exogenously expressed in mouse adult liver, fetal liver transcripts show higher expression and possess longer poly(A) tails, suggesting that IMP1 stabilizes them. IMP1 declines concomitantly with fetal liver transcripts as liver matures. Instead, RNA-binding proteins (RBPs) that promote RNA decay, such as cold shock domain containing protein E1 (CSDE1), K-homology domain splicing regulatory protein (KSRP), and CUG-BP1 and ETR3-like factors 1 (CELF1), bind to 3′ regions of fetal liver transcripts. These data suggest that transitions among RBPs associated with fetal liver transcripts shift regulation from stabilization to decay, leading to a postnatal decrease in those fetal transcripts. Full article

Post-transcriptional mechanisms play critical roles in the control of gene expression during neuronal development and maturation as they allow for faster responses to environmental cues and provide spatially-restricted compartments for local control of protein expression. These mechanisms depend on the interaction of cis-acting elements present in the mRNA sequence and trans-acting factors, such as RNA-binding proteins (RBPs) and microRNAs (miRNAs) that bind to those cis-elements and regulate mRNA stability, subcellular localization, and translation. Recent studies have uncovered an unexpected complexity in these interactions, where coding and non-coding RNAs, termed competing endogenous RNAs (ceRNAs), compete for binding to miRNAs. This competition can, thereby, control a larger number of miRNA target transcripts. However, competing RNA networks also extend to competition between target mRNAs for binding to limited amounts of RBPs. In this review, we present evidence that competitions between target mRNAs for binding to RBPs also occur in neurons, where they affect transcript stability and transport into axons and dendrites as well as translation. In addition, we illustrate the complexity of these mechanisms by demonstrating that RBPs and miRNAs also compete for target binding and regulation. Full article

The peptidyl-prolyl cis-trans isomerases (PPIases) that include immunophilins (cyclophilins and FKBPs) and parvulins (Pin1, Par14, Par17) participate in cell signaling, transcription, pre-mRNA processing and mRNA decay. The human genome encodes 19 cyclophilins, 18 FKBPs and three parvulins. Immunophilins are receptors for the immunosuppressive drugs cyclosporin A, FK506, and rapamycin that are used in organ transplantation. Pin1 has also been targeted in the treatment of Alzheimer’s disease, asthma, and a number of cancers. While these PPIases are characterized as molecular chaperones, they also act in a nonchaperone manner to promote protein-protein interactions using surfaces outside their active sites. The immunosuppressive drugs act by a gain-of-function mechanism by promoting protein-protein interactions in vivo. Several immunophilins have been identified as components of the spliceosome and are essential for alternative splicing. Pin1 plays roles in transcription and RNA processing by catalyzing conformational changes in the RNA Pol II C-terminal domain. Pin1 also binds several RNA binding proteins such as AUF1, KSRP, HuR, and SLBP that regulate mRNA decay by remodeling mRNP complexes. The functions of ribonucleoprotein associated PPIases are largely unknown. This review highlights PPIases that play roles in RNA-mediated gene expression, providing insight into their structures, functions and mechanisms of action in mRNP remodeling in vivo. Full article