Modulation of PKM alternative splicing by PTBP1 promotes gemcitabine resistance in pancreatic cancer cells

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive and incurable disease. Poor prognosis is due to multiple reasons, including acquisition of resistance to gemcitabine, the first-line chemotherapeutic approach. Thus, there is a strong need for novel therapies, targeting more directly the molecular aberrations of this disease. We found that chronic exposure of PDAC cells to gemcitabine selected a subpopulation of cells that are drug-resistant (DR-PDAC cells). Importantly, alternative splicing (AS) of the pyruvate kinase gene (PKM) was differentially modulated in DR-PDAC cells, resulting in promotion of the cancer-related PKM2 isoform, whose high expression also correlated with shorter recurrence-free survival in PDAC patients. Switching PKM splicing by antisense oligonucleotides to favor the alternative PKM1 variant rescued sensitivity of DR-PDAC cells to gemcitabine and cisplatin, suggesting that PKM2 expression is required to withstand drug-induced genotoxic stress. Mechanistically, upregulation of the polypyrimidine-tract binding protein (PTBP1), a key modulator of PKM splicing, correlated with PKM2 expression in DR-PDAC cell lines. PTBP1 was recruited more efficiently to PKM pre-mRNA in DR- than in parental PDAC cells. Accordingly, knockdown of PTBP1 in DR-PDAC cells reduced its recruitment to the PKM pre-mRNA, promoted splicing of the PKM1 variant and abolished drug resistance. Thus, chronic exposure to gemcitabine leads to upregulation of PTBP1 and modulation of PKM AS in PDAC cells, conferring resistance to the drug. These findings point to PKM2 and PTBP1 as new potential therapeutic targets to improve response of PDAC to chemotherapy.Oncogene advance online publication, 3 August 2015; doi:10.1038/onc.2015.270

Endocrinological applications in crustacean broodstock development- Winter School on Recent Advances in Breeding and Larviculture of Marine Finfish and Shellfish

Globally the scientific community is engaged in research on induced maturation of commercially important crustaceans aiming to the betterment of technologies over the already available eyestalk ablation techniques. In spite of all the technologies so far, there remains number of areas where further development would be highly desirable for optimizing commercial seed production

Embryonic stem cell-derived CD166+ precursors develop into fully functional sinoatrial-like cells

Rationale: A cell-based biological pacemaker is based on the differentiation of stem cells and the selection of a population displaying the molecular and functional properties of native sinoatrial node (SAN) cardiomyocytes. So far, such selection has been hampered by the lack of proper markers. CD166 is specifically but transiently expressed in the mouse heart tube and sinus venosus, the prospective SAN. Objective: We have explored the possibility of using CD166 expression for isolating SAN progenitors from differentiating embryonic stem cells. Methods and Results: We found that in embryonic day 10.5 mouse hearts, CD166 and HCN4, markers of the pacemaker tissue, are coexpressed. Sorting embryonic stem cells for CD166 expression at differentiation day 8 selects a population of pacemaker precursors. CD166(+) cells express high levels of genes involved in SAN development (Tbx18, Tbx3, Isl-1, Shox2) and function (Cx30.2, HCN4, HCN1, CaV1.3) and low levels of ventricular genes (Cx43, Kv4.2, HCN2, Nkx2.5). In culture, CD166(+) cells form an autorhythmic syncytium composed of cells morphologically similar to and with the electrophysiological properties of murine SAN myocytes. Isoproterenol increases (+57%) and acetylcholine decreases (-23%) the beating rate of CD166-selected cells, which express the -adrenergic and muscarinic receptors. In cocultures, CD166-selected cells are able to pace neonatal ventricular myocytes at a rate faster than their own. Furthermore, CD166(+) cells have lost pluripotency genes and do not form teratomas in vivo. Conclusions: We demonstrated for the first time the isolation of a nonteratogenic population of cardiac precursors able to mature and form a fully functional SAN-like tissue

The Stability of the Induced Epigenetic Programs

For many years scientists have been attracted to the possibility of changing cell identity. In the last decades seminal discoveries have shown that it is possible to reprogram somatic cells into pluripotent cells and even to transdifferentiate one cell type into another. In view of the potential applications that generating specific cell types in the laboratory can offer for cell-based therapies, the next important questions relate to the quality of the induced cell types. Importantly, epigenetic aberrations in reprogrammed cells have been correlated with defects in differentiation. Therefore, a look at the epigenome and understanding how different regulators can shape it appear fundamental to anticipate potential therapeutic pitfalls. This paper covers these epigenetic aspects in stem cells, differentiation, and reprogramming and discusses their importance for the safety of in vitro engineered cell types

Glycome dynamics in T and B cell development: basic immunological mechanisms and clinical applications

Glycans cover the surfaces of all mammalian cells through a process called glycosylation. Nearly all proteins and receptors that integrate the intricate series of co-stimulatory/inhibitory pathways of the immune system are glycosylated. Growing evidence indicates that the development of the immune system at the origins of T and B cell development is tightly regulated by glycosylation. In this opinion, we hypothesize that the glycome composition of developing T and B cells is developmentally regulated. We discuss how glycans play fundamental roles in lymphocyte development and how glycans early define T and B cell functionality in multiple aspects of adaptive immunity. These advances can provide opportunities for the discovery of novel disease factors and more effective candidate treatments for various conditions.This work was funded by the EU (ERC, GlycanSwitch, Grant Agreement N° 101071386). Views and opinions expressed are, however, those of the author(s) only and do not necessarily reflect those of the European Union or the European Research Council Executive Agency. Funded by the European Union (GlycanTrigger, Grant Agreement N°: 101093997. Views and opinions expressed are, however, those of the author(s) only and do not necessarily reflect those of the European Union or European Health and Digital Executive Agency. Neither the European Union nor the granting authority can be held responsible for them. M.M.V. and E.L-G. received funding from the Portuguese Foundation for Science and Technology (FCT) of the Portuguese Ministry of Science, Technology and Higher Education (M.M.V.: PD/BD/135452/2017, COVID/BD/152488/2022; E.L-G.: UI/BD/152866/2022)

Scd1 plays a tumor-suppressive role in survival of leukemia stem cells and the development of chronic myeloid leukemia

Chronic myeloid leukemia (CML) is derived from a stem cell, and it is widely accepted that the existence of leukemia stem cells (LSCs) is one of the major reasons for the relapse of CML treated with kinase inhibitors. Key to eradicating LSCs is to identify genes that play a critical role in survival regulation of these stem cells. Using BCR-ABL-induced CML mouse model, here we show that expression of the stearoyl-CoA desaturase 1 (Scd1) gene is downregulated in LSCs and that Scd1 plays a tumor-suppressive role in LSCs with no effect on the function of normal hematopoietic stem cells. Deletion of Scd1 causes acceleration of CML development and conversely overexpression of Scd1 delays CML development. In addition, using genetic approaches, we show that Pten, p53, and Bcl2 are regulated by Scd1 in LSCs. Furthermore, we find that induction of Scd1 expression by a PPARγ agonist suppresses LSCs and delays CML development. Our results demonstrate a critical role for Scd1 in functional regulation of LSCs, providing a new anti-LSC strategy through enhancing Scd1 activity

Fourier Transform Infrared Microscopy for Probing Changes in Biomolecular Composition of Prokaryotic and Eukaryotic Cells in Response to External Stressors

ABSTRACT Fourier Transform Infrared Spectroscopy (FTIR) is a tool that allows for the characterization of molecular changes in biological systems. In this thesis, FTIR Microscopy is used to investigate the innate response of cells, via their biomolecular compositions, to nutrient stress and drug-induced changes. FTIR was used to study cyanobacteria (Synechocystis sp. PCC 6803 and Spirulina platensis) and algal (Neochloris minuta and Neochloris alveolaris) cells. A strain of yeast (Yarrowia lipolytica), and prostate cancer cells from the PC3 cell line were also studied. The chemometric analysis, in particular, principal component analysis (PCA), was used to differentiate different cell types, cells at various stages of growth, and metabolic modifications in cells in response to external stress. Changes in the cell biomolecular composition indicate biomarkers that indicate how cells respond to changes in their environment. The cell is in effect a sentinel or biological sensor, which can provide information on drug efficacy, and even on pollutants and other metabolites in the environment in which cells grow, not to mention a variety of pathogenic diseased states of cells. FTIR spectra of cells provide information on the composition of lipids, carbohydrates, and proteins within the cell. These biomolecules absorb IR radiation for the most part in distinct spectral regions. C-H, N-H, and O-H stretching vibrations occur in the 3300-2800 cm-1 region. Proteins absorb around 1650 and 1550 cm-1. While lipids, including phospholipids, nucleic acids, and polysaccharides display absorption in the 1400-900 cm-1 region. To test and verify the level of applicability of the FTIR technique we introduce the hypothesis that cyanobacterial cells in various stages of growth can be distinguished based on their FTIR spectrum. Different cyanobacterial strains at similar growth stages can also be characterized. IR spectra of prostate cancer cells in the presence of various drugs were also described based on their FTIR spectra