The involvement of microRNAs (miRNAs) in orchestrating self-renewal and differentiation of stem cells has been revealed in a number of recent studies. And while in human pluripotent stem cells, miRNAs have been directly linked to the core pluripotency network, including the cell cycle regulation and the maintenance of the self-renewing capacity, their role in the onset of differentiation in other contexts, such as determination of neural cell fate, remains poorly described. To bridge this gap, we used three model cell types to study miRNA expression patterns: human embryonic stem cells (hESCs), hESCs-derived self-renewing neural stem cells (NSCs), and differentiating NSCs. The comprehensive miRNA profiling presented here reveals novel sets of miRNAs differentially expressed during human neural cell fate determination in vitro. Furthermore, we report a miRNA expression profile of self-renewing human NSCs, which has been lacking to this date. Our data also indicates that miRNA clusters enriched in NSCs share the target-determining seed sequence with cell cycle regulatory miRNAs expressed in pluripotent hESCs. Lastly, our mechanistic experiments confirmed that cluster miR-17-92, one of the NSCs-enriched clusters, is directly transcriptionally regulated by transcription factor c-MYC.

• Klíčová slova
• Cell cycle, Human pluripotent stem cells, Neural stem cells, miRNA sequencing, microRNA,
• MeSH
• buněčná diferenciace genetika   MeSH
• embryonální kmenové buňky MeSH
• lidé MeSH
• mikro RNA * genetika   metabolismus   MeSH
• nervové kmenové buňky * metabolismus   MeSH
• stanovení celkové genové exprese MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• mikro RNA * MeSH

Recirculation of chronic lymphocytic leukemia (CLL) cells between the peripheral blood and lymphoid niches plays a critical role in disease pathophysiology, and inhibiting this process is one of the major mechanisms of action for B-cell receptor (BCR) inhibitors such as ibrutinib and idelalisib. Migration is a complex process guided by chemokine receptors and integrins. However, it remains largely unknown how CLL cells integrate multiple migratory signals while balancing survival in the peripheral blood and the decision to return to immune niches. Our study provided evidence that CXCR4/CD5 intraclonal subpopulations can be used to study the regulation of migration of CLL cells. We performed RNA profiling of CXCR4dimCD5bright vs CXCR4brightCD5dim CLL cells and identified differential expression of dozens of molecules with a putative function in cell migration. GRB2-associated binding protein 1 (GAB1) positively regulated CLL cell homing capacity of CXCR4brightCD5dim cells. Gradual GAB1 accumulation in CLL cells outside immune niches was mediated by FoxO1-induced transcriptional GAB1 activation. Upregulation of GAB1 also played an important role in maintaining basal phosphatidylinositol 3-kinase (PI3K) activity and the "tonic" AKT phosphorylation required to sustain the survival of resting CLL B cells. This finding is important during ibrutinib therapy, because CLL cells induce the FoxO1-GAB1-pAKT axis, which represents an adaptation mechanism to the inability to home to immune niches. We have demonstrated that GAB1 can be targeted therapeutically by novel GAB1 inhibitors, alone or in combination with BTK inhibition. GAB1 inhibitors induce CLL cell apoptosis, impair cell migration, inhibit tonic or BCR-induced AKT phosphorylation, and block compensatory AKT activity during ibrutinib therapy.

• MeSH
• adaptorové proteiny signální transdukční biosyntéza   MeSH
• adenin analogy a deriváty   farmakologie   MeSH
• chronická lymfatická leukemie farmakoterapie   metabolismus   patologie   MeSH
• forkhead box protein O1 metabolismus   MeSH
• lidé MeSH
• nádorové buněčné linie MeSH
• piperidiny farmakologie   MeSH
• pohyb buněk * MeSH
• protoonkogenní proteiny c-akt metabolismus   MeSH
• regulace genové exprese u leukemie * MeSH
• signální transdukce * MeSH
• upregulace * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• adaptorové proteiny signální transdukční MeSH
• adenin MeSH
• forkhead box protein O1 MeSH
• FOXO1 protein, human MeSH Prohlížeč
• GAB1 protein, human MeSH Prohlížeč
• ibrutinib MeSH Prohlížeč
• piperidiny MeSH
• protoonkogenní proteiny c-akt MeSH

B-cell receptor (BCR) signaling and T-cell interactions play a pivotal role in chronic lymphocytic leukemia (CLL) pathogenesis and disease aggressiveness. CLL cells can use microRNAs (miRNAs) and their targets to modulate microenvironmental interactions in the lymph node niches. To identify miRNA expression changes in the CLL microenvironment, we performed complex profiling of short noncoding RNAs in this context by comparing CXCR4/CD5 intraclonal cell subpopulations (CXCR4dimCD5bright vs CXCR4brightCD5dim cells). This identified dozens of differentially expressed miRNAs, including several that have previously been shown to modulate BCR signaling (miR-155, miR-150, and miR-22) but also other candidates for a role in microenvironmental interactions. Notably, all 3 miR-29 family members (miR-29a, miR-29b, miR-29c) were consistently down-modulated in the immune niches, and lower miR-29(a/b/c) levels associated with an increased relative responsiveness of CLL cells to BCR ligation and significantly shorter overall survival of CLL patients. We identified tumor necrosis factor receptor-associated factor 4 (TRAF4) as a novel direct target of miR-29s and revealed that higher TRAF4 levels increase CLL responsiveness to CD40 activation and downstream nuclear factor-κB (NF-κB) signaling. In CLL, BCR represses miR-29 expression via MYC, allowing for concurrent TRAF4 upregulation and stronger CD40-NF-κB signaling. This regulatory loop is disrupted by BCR inhibitors (bruton tyrosine kinase [BTK] inhibitor ibrutinib or phosphatidylinositol 3-kinase [PI3K] inhibitor idelalisib). In summary, we showed for the first time that a miRNA-dependent mechanism acts to activate CD40 signaling/T-cell interactions in a CLL microenvironment and described a novel miR-29-TRAF4-CD40 signaling axis modulated by BCR activity.

• MeSH
• adenin analogy a deriváty   farmakologie   MeSH
• antigeny CD40 genetika   metabolismus   MeSH
• chronická lymfatická leukemie farmakoterapie   genetika   metabolismus   patologie   MeSH
• dospělí MeSH
• faktor 4 asociovaný s receptory TNF genetika   metabolismus   MeSH
• lidé středního věku MeSH
• lidé MeSH
• mikro RNA genetika   MeSH
• míra přežití MeSH
• nádorové biomarkery genetika   metabolismus   MeSH
• nádorové buňky kultivované MeSH
• následné studie MeSH
• piperidiny farmakologie   MeSH
• prognóza MeSH
• protoonkogenní proteiny c-bcr antagonisté a inhibitory   MeSH
• protoonkogenní proteiny c-myc genetika   metabolismus   MeSH
• regulace genové exprese u nádorů * MeSH
• senioři MeSH
• Check Tag
• dospělí MeSH
• lidé středního věku MeSH
• lidé MeSH
• mužské pohlaví MeSH
• senioři MeSH
• ženské pohlaví MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• adenin MeSH
• antigeny CD40 MeSH
• BCR protein, human MeSH Prohlížeč
• faktor 4 asociovaný s receptory TNF MeSH
• ibrutinib MeSH Prohlížeč
• mikro RNA MeSH
• MIRN29a microRNA, human MeSH Prohlížeč
• MYC protein, human MeSH Prohlížeč
• nádorové biomarkery MeSH
• piperidiny MeSH
• protoonkogenní proteiny c-bcr MeSH
• protoonkogenní proteiny c-myc MeSH
• TRAF4 protein, human MeSH Prohlížeč

The adaptive immune system is responsible for generating immunological response and immunological memory. Regulation of adaptive immunity including B cell and T cell biology was mainly understood from the protein and microRNA perspective. However, long non-coding RNAs (lncRNAs) are an emerging class of non-coding RNAs (ncRNAs) that influence key factors in lymphocyte biology such as NOTCH, PAX5, MYC and EZH2. LncRNAs were described to modulate lymphocyte activation by regulating pathways such as NFAT, NFκB, MYC, interferon and TCR/BCR signalling (NRON, NKILA, BCALM, GAS5, PVT1), and cell effector functions (IFNG-AS1, TH2-LCR). Here we review lncRNA involvement in adaptive immunity and the implications for autoimmune diseases (multiple sclerosis, inflammatory bowel disease, rheumatoid arthritis) and T/B cell leukaemias and lymphomas (CLL, MCL, DLBCL, T-ALL). It is becoming clear that lncRNAs are important in adaptive immune response and provide new insights into its orchestration.

• Klíčová slova
• Adaptive immunity, Autoimmune diseases, B cell, B/T cell activation, Leukaemia, LncRNAs, Lymphocyte development, Lymphoma, T cell, T cell polarization,
• MeSH
• adaptivní imunita genetika   MeSH
• aktivace lymfocytů genetika   MeSH
• lidé MeSH
• nemoc genetika   MeSH
• RNA dlouhá nekódující genetika   fyziologie   MeSH
• signální transdukce genetika   imunologie   MeSH
• T-lymfocyty fyziologie   MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• přehledy MeSH
• Názvy látek
• RNA dlouhá nekódující MeSH

The approval of BTK and PI3K inhibitors (ibrutinib, idelalisib) represents a revolution in the therapy of B cell malignancies such as chronic lymphocytic leukemia (CLL), mantle-cell lymphoma (MCL), diffuse large B cell lymphoma (DLBCL), follicular lymphoma (FL), or Waldenström's macroglobulinemia (WM). However, these "BCR inhibitors" function by interfering with B cell pathophysiology in a more complex way than anticipated, and resistance develops through multiple mechanisms. In ibrutinib treated patients, the most commonly described resistance-mechanism is a mutation in BTK itself, which prevents the covalent binding of ibrutinib, or a mutation in PLCG2, which acts to bypass the dependency on BTK at the BCR signalosome. However, additional genetic aberrations leading to resistance are being described (such as mutations in the CARD11, CCND1, BIRC3, TRAF2, TRAF3, TNFAIP3, loss of chromosomal region 6q or 8p, a gain of Toll-like receptor (TLR)/MYD88 signaling or gain of 2p chromosomal region). Furthermore, relative resistance to BTK inhibitors can be caused by non-genetic adaptive mechanisms leading to compensatory pro-survival pathway activation. For instance, PI3K/mTOR/Akt, NFkB and MAPK activation, BCL2, MYC, and XPO1 upregulation or PTEN downregulation lead to B cell survival despite BTK inhibition. Resistance could also arise from activating microenvironmental pathways such as chemokine or integrin signaling via CXCR4 or VLA4 upregulation, respectively. Defining these compensatory pro-survival mechanisms can help to develop novel therapeutic combinations of BTK inhibitors with other inhibitors (such as BH3-mimetic venetoclax, XPO1 inhibitor selinexor, mTOR, or MEK inhibitors). The mechanisms of resistance to PI3K inhibitors remain relatively unclear, but some studies point to MAPK signaling upregulation via both genetic and non-genetic changes, which could be co-targeted therapeutically. Alternatively, drugs mimicking the BTK/PI3K inhibition effect can be used to prevent adhesion and/or malignant B cell migration (chemokine and integrin inhibitors) or to block the pro-proliferative T cell signals in the microenvironment (such as IL4/STAT signaling inhibitors). Here we review the genetic and non-genetic mechanisms of resistance and adaptation to the first generation of BTK and PI3K inhibitors (ibrutinib and idelalisib, respectively), and discuss possible combinatorial therapeutic strategies to overcome resistance or to increase clinical efficacy.

• Klíčová slova
• B cell malignancies, B cell receptor, BCR inhibitor, adaptation, ibrutinib, resistance, targeted therapy,
• Publikační typ
• časopisecké články MeSH

Oct4-mediated reprogramming has recently become a novel tool for the generation of various cell types from differentiated somatic cells. Although molecular mechanisms underlying this process are unknown, it is well documented that cells over-expressing Oct4 undergo transition from differentiated state into plastic state. This transition is associated with the acquisition of stem cells properties leading to epigenetically "open" state that is permissive to cell fate switch upon external stimuli. In order to contribute to our understanding of molecular mechanisms driving this process, we characterised human fibroblasts over-expressing Oct4 and performed comprehensive small-RNAseq analysis. Our analyses revealed new interesting aspects of Oct4-mediated cell plasticity induction. Cells over-expressing Oct4 lose their cell identity demonstrated by down-regulation of fibroblast-specific genes and up-regulation of epithelial genes. Interestingly, this process is associated with microRNA expression profile that is similar to microRNA profiles typically found in pluripotent stem cells. We also provide extensive network of microRNA families and clusters allowing us to precisely determine the miRNAome associated with the acquisition of Oct4-induced transient plastic state. Our data expands current knowledge of microRNA and their implications in cell fate alterations and contributing to understanding molecular mechanisms underlying it.

• MeSH
• buněčné linie MeSH
• embryo savčí * MeSH
• fibroblasty cytologie   metabolismus   MeSH
• lidé MeSH
• mikro RNA * biosyntéza   genetika   MeSH
• oktamerní transkripční faktor 3 * biosyntéza   genetika   MeSH
• regulace genové exprese * MeSH
• techniky buněčného přeprogramování * MeSH
• Check Tag
• lidé MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• mikro RNA * MeSH
• oktamerní transkripční faktor 3 * MeSH
• POU5F1 protein, human MeSH Prohlížeč