Circular RNAs (circRNAs) are an emerging class of RNA molecules that have been linked to human diseases and important regulatory pathways. Their functional roles are still under investigation, often hampered by inefficient circRNA formation in and ex vivo . We generated an intron-mediated enhancement (IME) system that—in comparison to previously published methods—increases circRNA formation up to 5-fold. This strategy also revealed previously undetected translation of circRNA, e.g., circRtn4. Substantiated by Western blots and mass spectrometry we showed that in mammalian cells, translation of circRtn4 containing a potential “infinite” circular reading frame resulted in “monomers” and extended proteins, presumably “multimer” tandem repeats. In order to achieve high levels of circRNA formation and translation of other natural or recombinant circRNAs, we constructed a versatile circRNA expression vector—pCircRNA-DMo. We demonstrated the general applicability of this method by efficiently generating two additional circRNAs exhibiting high expression levels. The circRNA expression vector will be an important tool to investigate different aspects of circRNA biogenesis and to gain insights into mechanisms of circular RNA translation.

Ageing, a leading cause of the decline/deficits in human learning, memory, and cognitive abilities, is a major risk factor for age-associated neurodegenerative disorders such as Alzheimer’s disease. Emerging evidence suggests that epigenetics, an inheritable but reversible biochemical process, plays a crucial role in the pathogenesis of age-related neurological disorders. DNA methylation, the best-known epigenetic mark, has attracted most attention in this regard. DNA methyltransferases (DNMTs) are key enzymes in mediating the DNA methylation process, by which a methyl group is transferred, faithfully or anew, to genomic DNA sequences. Biologically, DNMTs are important for gene imprinting. Accumulating evidence suggests that DNMTs not only play critical roles, including gene imprinting and transcription regulation, in early development stages of the central nervous system (CNS), but also are indispensable in adult learning, memory, and cognition. Therefore, the impact of DNMTs and DNA methylation on age-associated cognitive functions and neurodegenerative diseases has emerged as a pivotal topic in the field. In this review, the effects of each DNMT on CNS development and healthy and pathological ageing are discussed.

Vitamin D is an essential fat-soluble vitamin with multiple functions. Vitamin D receptor has been shown to be expressed in several types of immune cells suggesting vitamin D may have immune regulatory roles. Vitamin D insufficiency has been suggested to increase the risk of autoimmune diseases. However, little is known regarding its immunomodulatory effects in the condition of immune suppression. The aim of the present study was to investigate the regulatory effects of vitamin D on immune function in immunosuppressant mice. An immunosuppressant mouse model was induced by intraperitoneal injection with glucocorticiod for 3 days. Immunosuppressant mice were intragastrically administered with 1,25-dihydroxy-vitamin D3 [1,25(OH)D; 0,4, 6 or 10 IU/g body weight] for 7 days. On day 8, the mice were decapitated. The body weight and the weights of thymus and spleen were measured. Thymus and spleen indexes were calculated. The ratio of CD4/CD8 T lymphocytes in the peripheral blood, proliferation and interleukin-2 (IL-2) production of spleen T lymphocytes was detected. Compared with the mice in the control group, the body weight, thymus and spleen indexes, the ratios of CD4/CD8 in peripheral blood and IL-2 production and proliferation of spleen T lymphocytes were decreased in immunosuppressant mice induced by glucocorticiod. However, in vitamin D-treated mice, the thymus indexes, the ratios of CD4/CD8, secretion of IL-2 and the proliferation index of spleen T lymphocytes were significantly increased (P<0.05). Among the three doses of 1,25(OH)D, 6 IU/g was most effective in improving the immune function. These results indicate that vitamin D supplementation can improve immune recovery in immunosuppressant mice by stimulating T-cell proliferation and elevating IL-2 production.

Dnmt3a2, a de novo DNA methyltransferase, is induced by neuronal activity and participates in long-term memory formation with the increased expression of synaptic plasticity genes. We wanted to determine if Dnmt3a2 with its partner Dnmt3L may influence motor behavior via the dopaminergic system. To this end, we generated a mouse line, Dnmt3a2/3LDat/wt, with dopamine transporter (DAT) promotor driven Dnmt3a2/3L overexpression. The mice were studied with behavioral paradigms (e.g., cylinder test, open field, and treadmill), brain slice patch clamp recordings, ex vivo metabolite analysis, and in vivo positron emission tomography (PET) using the dopaminergic tracer 6-[18F]FMT. The results showed that spontaneous activity and exercise performance were enhanced in Dnmt3a2/3LDat/wt mice compared to Dnmt3a2/3Lwt/wt controls. Dopaminergic substantia nigra pars compacta neurons of Dnmt3a2/3LDat/wt animals displayed a higher fire frequency and excitability. However, dopamine concentration was not increased in the striatum, and dopamine metabolite concentration was even significantly decreased. Striatal 6-[18F]FMT uptake, reflecting aromatic L-amino acid decarboxylase activity, was the same in Dnmt3a2/3LDat/wt mice and controls. [18F]FDG PET showed that hypothalamic metabolic activity was tightly linked to motor behavior in Dnmt3a2/3LDat/wt mice. Furthermore, dopamine biosynthesis and motor-related metabolic activity were correlated in the hypothalamus. Our findings suggest that Dnmt3a2/3L, when overexpressed in dopaminergic neurons, modulates motor performance via activation of the nigrostriatal pathway. This does not involve increased dopamine synthesis.

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