miRNA

miRNA: Fine tuning the levels of mRNA and proteins

The cellular processes are controlled by a narrow range ofmRNA and proteins levels, where small RNAs (sRNAs) known asmiRNAs play a critical role. miRNA plays an essential role in modulating gene expression bybinding to complementary sites on the target mRNA, thereby blockingtranslation or degrading the target mRNA. The regulation of miRNA function, spatial localization of pre- and mature miRNA is not understood. Our lab is interest in the following areas:

A. Spatio-temporal dynamics of miRNA

The main focus of lab is spatial regulation of miRNA and its dynamic association with mitochondria in human cell.

The pri-miRNA is transcribed from genome by RNA polymerase II. The pri-miRNA is processed into pre-miRNA by RNase III enzyme Dorsha with the help of its partner Pasha. The 3′ nt overhang is recognised by Exportin-5 which exports the pre-miRNA into the cytosol in Ran-GTP dependent manner. In cytosol, pri-miRNA is cleaved by Dicer to produce mature miRNA which then becomes a part of RISC assembly to attenuate or degrade complimentary mRNA. pre-miRNA and mature miRNA are translocated to various subcellular locations (nucleus, ER, P-bodies, etc.) including mitochondria.

We were one of the pioneers in discussing the association of pre-miRNA, mature miRNA and core component of RISC, Ago2/3 with mitochondria. The store house of miRNA, P-bodies is also associated with mitochondria.

The emerging evidence and current understanding of the field arises interesting aspects like;

1. Mitochondrial outer membrane may itself serve as a novel platform for the transport or assembly miRNA and associated components to fine tune local protein synthesis at specific foci in the cell.

2. The association and translocation of miRNA at mitochondria is dynamic process and stimuli dependent.

3. miRNA may regulate the mitochondrial genome and consequently its functions and cellular processes.

4. Mitochondria may protect and preserve miRNA for their own regulation and overall cellular signalling.

5. miRNAmay mediate massive nuclear-mitochondrial crosstalk.

6. The association may be sensitive to physiologocial and pathological cues

B. miRNA association dynamics in Fragile X Mental retardation syndrome

Fragile X-associated Tremor/Ataxia Syndrome (FXTAS) is a X-linked inheritedneurodegenerative disorder that affects older (>50 years) adult males who are carriers of an expansionof 55 to 200 CGG repeats in the 5’-untranslated region of the Fragile X Mental Retardation 1 (FMR1)gene, which is also referred to “premutation allele”.

The clinical features of FXTAS include progressive development of tremor, ataxia andneuropsychological problems, including parkinsonism, peripheral neuropathy, memory impairment, dementia and cognitive decline.

The prevalence of premutation carriers of expanded CGG repeats in the general population isestimated to be 1 in 250 females and ~1 in 400 males, resulting with incomplete penetrance, to aprevalence of 1 in ~3,000 males that may develop clinical features of FXTAS. Therefore, FXTAS isone of the most common single gene causes of tremor, ataxia and cognitive decline among older adults. Females may also present with FXTAS, although less commonly comparedto males, presumably because of the protective effect of the second X chromosome.Principal neuropathology of FXTAS includes mild brain atrophy and white matter lesions, withthe presence of ubiquitin-positive nuclear neuronal and astrocytic inclusions.

The striking characteristic of FXTAS is that, in contrast to Fragile X syndrome, where fullmutations (>200 CGG repeats) result in hyper-methylation and silencing of the FMR1 gene, FXTASpatients carrier of shorter CGG expansions (55–200 CGG repeats) present increased expression ofFMR1 mRNA levels and normal, or near normal, FMRP protein expression.

The remarkable feature is the ubiquitin-positive neuronal aggregates, which are characteristic of FXTAS, contain the pathogenic mutant FMR1 RNA. The RNA aggregatesequesters RNA binding proteins, DROSHA and DGCR8 within. This results in miRNAs processing defects and ultimately in decreased expression ofmature miRNAs. Importantly, the sole overexpression of DGCR8 restored to normal both the dendriticmorphological abnormalities and the loss of neuronal cells, demonstrating that titration of DGCR8 byexpanded CGG repeats is a leading eventually to neuronal cell death.

The poly-G protein aggregates sequesters RNA binding proteins, some mitochondrial proteins which are involved in various cellular processes including cell metabolism, cell death and neuronal functions.

The emerging evidence and current understanding of the field arises interesting aspects like;

1. The RNA aggregates may affect the site specific localisation of Ago2/3 and miRNAs at mitochondria dynamically.

2. The poly-G aggregates may form defective puncta and imbibe affinity based mitochondrial component proteins causing defects in mitochondrial functions.

3. The repeats may disrupt the site specific miRNA mediated fine tuning at mitochondria and its dynamics

C. miRNA dynamics in Parkinson’s Disease and miRNA mediated rescue in defective parkinsonian autophagy

Parkinson’s disease (PD) is second most frequent neurodegenerative disease affecting individuals over 50 years of age. The symptoms are bradykinesia, resting tremor and muscle tone rigidity. Onset of symptoms is the result of loss of neurons substantia nigra pars compacta (SNpc). The hallmark of PD is occurrence of Lewy bodies (LB) which are composed of α-synuclein (α-Syn) and ubiquitinated proteins in extant dopaminergic neurons. LB spread in other non-dopaminergic areas of the brain including brainstem cells nuclei, olfactory bulb, limbic system and neocortex suggesting cell-to-cell propagation of α-Syn or other regulators.

Our focus is to;

1. To identify the altered miRNAs in parkinsonian stress.

2. The modulation of miRNA levels to rescue mitochondrial functions.

3. To rescue miRNA mediated defective autophagy.

In future we are planning to understand

1. The dynamics of exosome mediated propagation of these pathologies.

2. To identify the delivery of miRNA via exosomes

3. To identify the effect of exosomal miRNA on mitochondria

4. The role and functional alterations of mitochondria and the involvement of exosomal miRNA.