35 research outputs found
miR-34a-5p as molecular hub of pathomechanisms in Huntington's disease
Background Although a pivotal role of microRNA (miRNA, miR) in the pathogenesis of Huntingtonâs disease (HD)
is increasingly recognized, the molecular functions of miRNAs in the pathomechanisms of HD await further elucidation. One of the miRNAs that have been associated with HD is miR-34a-5p, which was deregulated in the mouse R6/2
model and in human HD brain tissues.
Methods The aim of our study was to demonstrate interactions between miR-34a-5p and HD associated genes.
By computational means we predicted 12 801 potential target genes of miR-34a-5p. An in-silico pathway analysis
revealed 22 potential miR-34a-5p target genes in the KEGG (Kyoto Encyclopedia of Genes and Genomes) pathway
âHuntingtonâs diseaseâ.
Results Using our high-throughput miRNA interaction reporter assay (HiTmIR) we identifed NDUFA9, TAF4B, NRF1,
POLR2J2, DNALI1, HIP1, TGM2 and POLR2G as direct miR-34a-5p target genes. Direct binding of miR-34a-5p to target
sites in the 3âUTRs of TAF4B, NDUFA9, HIP1 and NRF1 was verifed by a mutagenesis HiTmIR assay and by determining
endogenous protein levels for HIP1 and NDUFA9. STRING (Search Tool for the Retrieval of Interacting Genes/Proteins)
analysis identifed proteinâprotein interaction networks associated with HD like âGlutamine Receptor Signaling Pathwayâ and âCalcium Ion Transmembrane Import Into Cytosolâ.
Conclusion Our study demonstrates multiple interactions between miR-34a-5p and HD associated target genes and
thereby lays the ground for future therapeutic interventions using this miRNA
Reduced prevalence of placental malaria in primiparae with blood group O
Background Blood group O protects African children against severe malaria and
has reached high prevalence in malarious regions. However, its role in malaria
in pregnancy is ambiguous. In 839 delivering Ghanaian women, associations of
ABO blood groups with Plasmodium falciparum infection were examined. Methods
Plasmodium falciparum infection was diagnosed in placental blood samples by
microscopy and PCR assays. Present or past infection was defined as the
detection of parasitaemia or haemozoin by microscopy, or a positive PCR
result. Blood groups were inferred from genotyping rs8176719 (indicating the O
allele) and rs8176746/rs8176747 (distinguishing the B allele from the A
allele). Results The majority of women had blood group O (55.4%); present or
past P. falciparum infection was seen in 62.3% of all women. Among multiparae,
the blood groups had no influence on P. falciparum infection. In contrast,
primiparae with blood group O had significantly less present or past infection
than women with non-O blood groups (61.5 vs 76.2%, Pâ=â0.007). In multivariate
analysis, the odds of present or past placental P. falciparum infection were
reduced by 45% in blood group O primiparae (aOR, 0.55 [95% CI, 0.33â0.94]).
Conclusions The present study shows a clear protective effect of blood group O
against malaria in primiparae. This accords with findings in severe malaria
and in vitro results. The data underline the relevance of host genetic
protection among primiparae, i.e. the high-risk group for malaria in
pregnancy, and contribute to the understanding of high O allele frequencies in
Africa
Induction of the Endoplasmic-Reticulum-Stress Response: MicroRNA-34a Targeting of the IRE1α-Branch
Neurodegenerative disorders such as Alzheimerâs disease (AD) and Parkinsonâs disease
(PD) are characterized by the accumulation of misfolded proteins in the endoplasmic reticulum (ER)
and the unfolded protein response (UPR). Modulating the UPR is one of the major challenges to
counteract the development of neurodegenerative disorders and other diseases with affected UPR.
Here, we show that miR-34a-5p directly targets the IRE1α branch of the UPR, including the genes BIP,
IRE1α, and XBP1. Upon induction of ER stress in neuronal cells, miR-34a-5p overexpression impacts
the resulting UPR via a significant reduction in IRE1α and XBP1s that in turn leads to decreased
viability, increased cytotoxicity and caspase activity. The possibility to modify the UPR signaling
pathway by a single miRNA that targets central genes of the IRE1α branch offers new perspectives
for future therapeutic approaches against neurodegeneration
miR-34a as hub of T cell regulation networks
Background: Micro(mi)RNAs are increasingly recognized as central regulators of immune cell function. While it has
been predicted that miRNAs have multiple targets, the majority of these predictions still await experimental confirmation.
Here, miR-34a, a well-known tumor suppressor, is analyzed for targeting genes involved in immune system processes of
leucocytes.
Methods: Using an in-silico approach, we combined miRNA target prediction with GeneTrail2, a web tool for Multi-omics
enrichment analysis, to identify miR-34a target genes, which are involved in the immune system process subcategory of
Gene Ontology.
Results: Out of the 193 predicted target genes in this subcategory we experimentally tested 22 target genes and
confirmed binding of miR-34a to 14 target genes including VAMP2, IKBKE, MYH9, MARCH8, KLRK1, CD11A, TRAFD1, CCR1,
PYDC1, PRF1, PIK3R2, PIK3CD, AP1B1, and ADAM10 by dual luciferase assays. By transfecting Jurkat, primary CD4+ and CD8+
T cells with miR-34a, we demonstrated that ectopic expression of miR-34a leads to reduced levels of endogenous VAMP2
and CD11A, which are central to the analyzed subcategories. Functional downstream analysis of miR-34a over-expression
in activated CD8+ T cells exhibits a distinct decrease of PRF1 secretion.
Conclusions: By simultaneous targeting of 14 mRNAs miR-34a acts as major hub of T cell regulatory networks suggesting
to utilize miR-34a as target of intervention towards a modulation of the immune responsiveness of T-cells in a broad
tumor context
Wrinkle in the plan: miR-34a-5p impacts chemokine signaling by modulating CXCL10/CXCL11/CXCR3-axis in CD4+, CD8+ T cells, and M1 macrophages
Background In 2016 the first-in-human phase I study of a miRNA-based cancer therapy with a liposomal mimic of microRNA-34a-5p (miR-34a-5p) was closed due to five immune related serious adverse events (SAEs) resulting in four patient deaths. For future applications of miRNA mimics in cancer therapy it is mandatory to unravel the miRNA effects both on the tumor tissue and on immune cells. Here, we set out to analyze the impact of miR-34a-5p over-expression on the CXCL10/CXCL11/CXCR3 axis, which is central for the development of an effective cancer control.
Methods We performed a whole genome expression analysis of miR-34a-5p transfected M1 macrophages followed by an over-representation and a proteinâprotein network analysis. In-silico miRNA target prediction and dual luciferase assays were used for target identification and verification. Target genes involved in chemokine signaling were functionally analyzed in M1 macrophages, CD4+ and CD8+ T cells.
Results A whole genome expression analysis of M1 macrophages with induced miR-34a-5p over-expression revealed an interaction network of downregulated target mRNAs including CXCL10 and CXCL11. In-silico target prediction in combination with dual luciferase assays identified direct binding of miR-34a-5p to the 3âČUTRs of CXCL10 and CXCL11. Decreased CXCL10 and CXCL11 secretion was shown on the endogenous protein level and in the supernatant of miR-34a-5p transfected and activated M1 macrophages. To complete the analysis of the CXCL10/CXCL11/CXCR3 axis, we activated miR-34a-5p transfected CD4+ and CD8+ T cells by PMA/Ionomycin and found reduced levels of endogenous CXCR3 and CXCR3 on the cell surface.
Conclusions MiR-34a-5p mimic administered by intravenous administration will likely not only be up-taken by the tumor cells but also by the immune cells. Our results indicate that miR-34a-5p over-expression leads in M1 macrophages to a reduced secretion of CXCL10 and CXCL11 chemokines and in CD4+ and CD8+ T cells to a reduced expression of CXCR3. As a result, less immune cells will be attracted to the tumor site. Furthermore, high levels of miR-34a-5p in naive CD4+ T cells can in turn hinder Th1 cell polarization through the downregulation of CXCR3 leading to a less pronounced activation of cytotoxic T lymphocytes, natural killer, and natural killer T cells and possibly contributing to lymphocytopenia
miR-34a: a new player in the regulation of T cell function by modulation of NF-ÎșB signaling
NF-ÎșB functions as modulator of T cell receptor-mediated signaling and transcriptional regulator of miR-34a. Our in
silico analysis revealed that miR-34a impacts the NF-ÎșB signalosome with miR-34a binding sites in 14 key members of
the NF-ÎșB signaling pathway. Functional analysis identified five target genes of miR-34a including PLCG1, CD3E, PIK3CB,
TAB2, and NFÎBIA. Overexpression of miR-34a in CD4+ and CD8+ T cells led to a significant decrease of NFÎBIA as the
most downstream cytoplasmic NF-ÎșB member, a reduced cell surface abundance of TCRA and CD3E, and to a
reduction of T cell killing capacity. Inhibition of miR-34a caused an increase of NFÎBIA, TCRA, and CD3E. Notably,
activation of CD4+ and CD8+ T cells entrails a gradual increase of miR-34a. Our results lend further support to a model
with miR-34a as a central NF-ÎșB regulator in T cells
Modulation of intracellular calcium signaling by microRNA-34a-5p
Adjusting intracellular calcium signaling is an important feature in the regulation of immune cell function and survival. Here we show that miR-34a-5p, a small non-coding RNA that is deregulated in many common diseases, is a regulator of store-operated Ca2+ entry (SOCE) and calcineurin signaling. Upon miR-34a-5p overexpression, we observed both a decreased depletion of ER calcium content and a decreased Ca2+ influx through Ca2+ release-activated Ca2+ channels. Based on an in silico target prediction we identified multiple miR-34a-5p target genes within both pathways that are implicated in the balance between T-cell activation and apoptosis including ITPR2, CAMLG, STIM1, ORAI3, RCAN1, PPP3R1, and NFATC4. Functional analysis revealed a decrease in Ca2+ activated calcineurin pathway activity measured by a reduced IL-2 secretion due to miR-34a-5p overexpression. Impacting SOCE and/or downstream calcineurin/NFAT signaling by miR-34a-5p offers a possible future approach to manipulate immune cells for clinical interventions
Quantitative and time-resolved miRNA pattern of early human T cell activation
T cells are central to the immune response against
various pathogens and cancer cells. Complex networks of transcriptional and post-transcriptional
regulators, including microRNAs (miRNAs), coordinate the T cell activation process. Available miRNA
datasets, however, do not sufficiently dissolve the
dynamic changes of miRNA controlled networks
upon T cell activation. Here, we established a quantitative and time-resolved expression pattern for the
entire miRNome over a period of 24 h upon human Tcell activation. Based on our time-resolved datasets,
we identified central miRNAs and specified common miRNA expression profiles. We found the most
prominent quantitative expression changes for miR155-5p with a range from initially 40 molecules/cell to
1600 molecules/cell upon T-cell activation. We established a comprehensive dynamic regulatory network
of both the up- and downstream regulation of miR155. Upstream, we highlight IRF4 and its complexes
with SPI1 and BATF as central for the transcriptional
regulation of miR-155. Downstream of miR-155-5p,
we verified 17 of its target genes by the time-resolved
data recorded after T cell activation. Our data provide comprehensive insights into the range of stimulus induced miRNA abundance changes and lay the
ground to identify efficient points of intervention for
modifying the T cell response
miRNAs and sports: tracking training status and potentially confounding diagnoses
Background: The dependency of miRNA abundance from physiological processes such as exercises remains partially understood. We set out to analyze the effect of physical exercises on miRNA profiles in blood and plasma of endurance and strength athletes in a systematic manner and correlated differentially abundant miRNAs in athletes to disease miRNAs biomarkers towards a better understanding of how physical exercise may confound disease diagnosis by miRNAs. Methods: We profiled blood and plasma of 29 athletes before and after exercise. With four samples analyzed for each individual we analyzed 116 full miRNomes. The study set-up enabled paired analyses of individuals. Affected miRNAs were investigated for known disease associations using network analysis. Results: MiRNA patterns in blood and plasma of endurance and strength athletes vary significantly with differences in blood outreaching variations in plasma. We found only moderate differences between the miRNA levels before training and the RNA levels after training as compared to the more obvious variations found between strength athletes and endurance athletes. We observed significant variations in the abundance of miR-140-3p that is a known circulating disease markers (raw and adjusted p value of 5Â ĂÂ 10â12 and 4Â ĂÂ 10â7). Similarly, the levels of miR-140-5p and miR-650, both of which have been reported as makers for a wide range of human pathologies significantly depend on the training mode. Among the most affected disease categories we found acute myocardial infarction. MiRNAs, which are up-regulated in endurance athletes inhibit VEGFA as shown by systems biology analysis of experimentally validated target genes. Conclusion: We provide evidence that the mode and the extent of training are important confounding factors for a miRNA based disease diagnosis
Chemoradiotherapy-induced increase in Th17 cell frequency in cervical cancer patients is associated with therapy resistance and early relapse
Cervical cancer therapy is still a major clinical challenge, as patients substantially differ in their response to standard treatments, including chemoradiotherapy (CRT). During cervical carcinogenesis, T-helper (Th)-17 cells accumulate in the peripheral blood and tumor tissues of cancer patients and are associated with poor prognosis. In this prospective study, we find increased Th17 frequencies in the blood of patients after chemoradiotherapy and a post-therapeutic ratio of Th17/CD4+ T cells >â8% was associated with early recurrence. Furthermore, Th17 cells promote resistance of cervical cancer cells toward CRT, which was dependent on the AKT signaling pathway. Consistently, patients with high Th17 frequencies in pretherapeutic biopsies exhibit lower response to primary CRT. This work reveals a key role of Th17 cells in CRT resistance and elevated Th17 frequencies in the blood after CRT correspond with early recurrence. Our results may help to explain individual treatment responses of cervical cancer patients and suggest evaluation of Th17 cells as a novel predictive biomarker for chemoradiotherapy responses and as a potential target for immunotherapy in cervical cancer