Chronic metabolic and neurodegenerative syndromes such as obesity or Parkinson’s disease, represent a growing epidemic especially in the constantly aging populations. They result from a failure in the interplay of environmental factors, humoral signals, extra- and intracellular enzymatic cascades, genetic and epigenetic signals in the central nervous system and in the periphery. Physiological and pathophysiological mechanisms involved in this regulation require intensive research to develop novel therapeutic strategies.
Our research group has a strong interest in studying the role of distinct neuronal population-specific non-coding RNAs in chronic complex trait disorders. In perspective, the integration of expression and functional data from mouse and patient brain samples will provide novel insights for future genome wide association studies aiming to identify genetic variants within non-coding RNA genes and/or within the genes influenced by these RNA species.
Can we treat neurodegenerative diseases by preventing an age-related decline in microRNA expression?
Vinnikov, I.A., Domanskyi, A.
2017. Neural Regeneration Research 12, 1602-1604.
Abstract: MicroRNA pathway is down-regulated in aged dopaminergic neurons: Parkinson’s disease (PD) is the most frequent motor neurodegenerative disorder and is morphologically mainly associated with progressive dopaminergic neuronal loss in the ventral midbrain. The cause of this pathology is unknown, but aging is well established as the strongest risk factor, which by far prevails over gender, environmental and genetic factors. In our recent work (Chmielarz et al., 2017), we have demonstrated that the expression of Dicer, a multidomain ribonuclease III and a key endonuclease in microRNA (miRNA) maturation pathway, is significantly down-regulated in aged mouse midbrain. Further, using a laser-assisted microdissection and quantitative PCR profiling techniques, we analyzed microRNAomes of dopaminergic neurons from the mouse substantia nigra and identified a predominant decrease of miRNA expression in aged dopaminergic neurons. Dicer mRNA levels are also reduced in the ventral midbrain of PD patients (Simunovic et al., 2010). Importantly, several miRNAs have been shown to regulate PD-associated genes, suggesting that age-related deregulation of miRNA signaling may contribute to neurodegeneration (Heman-Ackah et al., 2013).
The two major questions arising from these observations are:
Does the miRNA-mediated regulation provide an essential protection mechanism from neurodegeneration?
May this protective component be compromised during aging and make the dopamine system more susceptible for other genetic and environmental factors leading to PD?
In our studies, we attempt to resolve these two challenges. Recently, we have found a way to answer the first of these two major questions: miRNA pathway indeed turned out to be cytoprotective for adult dopaminergic neurons (Chmielarz et al., 2017). An increasing number of published and ongoing studies also start to address the second question, identifying neuronal functions- and survival-regulating genes and pathways targeted by miRNA network in the context of neurodegeneration (Briggs et al., 2015).
Dicer and microRNAs protect adult dopamine neurons
Chmielarz P., Konovalova J., Najam S.S., Alter H., Piepponen T.P., Erfle H., Sonntag K.C., Schütz G., Vinnikov I.A., Domanskyi A.
2017. Cell Death and Disease 8(5): e2813.
Abstract: MicroRNAs (miRs) are important post-transcriptional regulators of gene expression implicated in neuronal development, differentiation, aging and neurodegenerative diseases, including Parkinson’s disease (PD). Several miRs have been linked to PD-associated genes, apoptosis and stress response pathways, suggesting that deregulation of miRs may contribute to the development of the neurodegenerative phenotype. Here, we investigate the cell-autonomous role of miR processing RNAse Dicer in the functional maintenance of adult dopamine (DA) neurons. We demonstrate a reduction of Dicer in the ventral midbrain and altered miR expression profiles in laser-microdissected DA neurons of aged mice. Using a mouse line expressing tamoxifen-inducible CreERT2 recombinase under control of the DA transporter promoter, we show that a tissue-specific conditional ablation of Dicer in DA neurons of adult mice led to decreased levels of striatal DA and its metabolites without a reduction in neuronal body numbers in hemizygous mice (DicerHET) and to progressive loss of DA neurons with severe locomotor deficits in nullizygous mice (DicerCKO). Moreover, we show that pharmacological stimulation of miR biosynthesis promoted survival of cultured DA neurons and reduced their vulnerability to thapsigargin-induced endoplasmic reticulum stress. Our data demonstrate that Dicer is crucial for maintenance of adult DA neurons, whereas a stimulation of miR production can promote neuronal survival, which may have direct implications for PD treatment.
Continuous Delivery of Oligonucleotides into the Brain
Vinnikov, I.A., Domanskyi, A., and Konopka, W.
2016. physiology (Neuromethods), Humana Press. 7: 9.
Abstract: The growing field of RNA neurobiology dictates development and improvement of effective and reliable in vivo techniques to address the function of particular microRNA molecules within the brain. Here we describe a novel method involving continuous delivery of oligonucleotides into a brain region of interest by osmotic pump infusion. The approach implements application of double-stranded microRNA-mimics with only two LNA moieties at the 30-end and additionally one at the 50-end of the sense strand. This method holds promise for long-lasting and specific siRNA upregulation in vivo, especially in the Dicer-depleted systems, where other approaches are limited or not applicable. Being robust and effective, various techniques described in this chapter can be easily modified in order to achieve up- or downregulation of expression of specific RNA molecules, bi- or unilateral infusions or injections, and in vivo “screening” strategy allowing to start from a bigger group of RNA molecules and end up with identification of single RNA species critical for a phenotype.
Transcription factors Foxa1 and Foxa2 are required for adult dopamine neurons maintenance
Domanskyi, A., Alter, H., Vogt, M.A., Gass, P. and Vinnikov, I.A.
2014. Frontiers in Cellular Neuroscience 8: 275.
Abstract: The proteins Foxa1 and Foxa2 belong to the forkhead family of transcription factors and are involved in the development of several tissues, including liver, pancreas, lung, prostate, and the neural system. Both Foxa1 and Foxa2 are also crucial for the specification and differentiation of dopamine (DA) neurons during embryonic development, while about 30% of mice with an embryonic deletion of a single allele of the Foxa2 gene exhibit an age-related asymmetric loss of DA neurons and develop locomotor symptoms resembling Parkinson’s disease (PD). Notably, both Foxa1 and Foxa2 factors continue to be expressed in the adult dopamine system. To directly assess their functions selectively in adult DA neurons, we induced genetic deletions of Foxa1/2 transcription factors in mice using a tamoxifen inducible tissue-specific CreERT2 recombinase expressed under control of the dopamine transporter (DAT) promoter (DATCreERT2). The conditional DA neurons-specific ablation of both genes, but not of Foxa2 alone, in early adulthood, caused a decline of striatal dopamine and its metabolites, along with locomotor deficits. At early pre-symptomatic stages, we observed a decline in aldehyde dehydrogenase family 1, subfamily A1 (Aldh1a1) protein expression in DA neurons. Further analyses revealed a decline of aromatic amino acid decarboxylase (AADC) and a complete loss of DAT expression in these neurons. These molecular changes ultimately led to a reduction of DA neuron numbers in the substantia nigra pars compacta (SNpc) of aged cFoxa1/2−/− mice, resembling the progressive course of PD in humans. Altogether, in this study, we address the molecular, cellular, and functional role of both Foxa1 and Foxa2 factors in the maintenance of the adult dopamine system which may help to find better approaches for PD treatment.
Hypothalamic miR-103 Protects from Hyperphagic Obesity in Mice
Vinnikov, I.A., Hajdukiewicz, K., Reymann, J., Beneke, J., Czajkowski, R., Roth, L.C., Novak, M., Roller, A., Dörner, N., Starkuviene, V., Theis, F.J., Erfle, H., Schütz, G., Grinevich, V. and Konopka, W.
2014. The Journal of Neuroscience 34(32): 10659-10674.
Abstract: The role of neuronal non-coding RNAs in energy control of the body is not fully understood. The arcuate nucleus (ARC) of the hypothalamus comprises neurons regulating food intake and body weight. Here we show that Dicer-dependent loss of microRNAs in these neurons of adult (DicerCKO) mice causes chronic overactivation of the signaling pathways involving phosphatidylinositol-3-kinase (PI3K), Akt, and mammalian target of rapamycin (mTOR) and an imbalance in the levels of neuropeptides, resulting in severe hyperphagic obesity. Similarly, the activation of PI3K-Akt-mTOR pathway due to Pten deletion in the adult forebrain leads to comparable weight increase. Conversely, the mTORC1 inhibitor rapamycin normalizes obesity in mice with an inactivated Dicer1 or Pten gene. Importantly, the continuous delivery of oligonucleotides mimicking microRNAs, which are predicted to target PI3K-Akt-mTOR pathway components, to the hypothalamus attenuates adiposity in DicerCKO mice. Furthermore, loss of miR-103 causes strong upregulation of the PI3K-Akt-mTOR pathway in vitro and its application into the ARC of the Dicer-deficient mice both reverses upregulation of Pik3cg, the mRNA encoding the catalytic subunit p110γ of the PI3K complex, and attenuates the hyperphagic obesity. Our data demonstrate in vivo the crucial role of neuronal microRNAs in the control of energy homeostasis.
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