Controlling molecular signatures at nanoscale level in live cells using light

Dr. Meenakshi Tanwar’s interview with Bio Patrika hosting “Vigyan Patrika”, a series of author interviews. Dr. Tanwar is currently a Post-Doctoral Fellow in the laboratory of Dr. Mini Jose and Prof. Deepak Nair at the Centre of Neuroscience, Indian Institute of Science, Bangalore. She completed her doctoral study in Biochemistry from the University of Delhi South Campus, New Delhi, under the supervision of Prof. Suneel Kateriya. Her research works focus on the development of optogenetic tools for modulation of signaling cascades and utilizing these tools to modulate and study the dynamics of cyclic nucleotide signaling and its downstream effector molecules at the nanoscale level in neurons. Her study helps to get a deeper insight into the dynamics and nanoscale organization of signaling molecules in a spatio-temporal manner. Google Scholar Link: https://scholar.google.com/citations?user=b7SlOJ8AAAAJ&hl=en. Here, Meenakshi talks about her work on “Optogenetic modulation of real-time nanoscale dynamics of HCN channels using photoactivated adenylyl cyclases” published in RSC Chem Biol (2021).

Listen audio

How would you explain your paper’s key results to the non-scientific community?

A cell is a complex system with numerous biomolecular complexes to regulate and perform cellular functions. Regulation of cellular functions is attained through the modulation and coordination of many signaling pathways. Cyclic nucleotide, a ubiquitous secondary messenger, regulates a diverse range of functions in gene expression, cellular proliferation and apoptosis, neuronal signaling, and microbial pathogenesis. cAMP also modulates the activity of Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels. HCN channels play a crucial role in maintaining the membrane potential and are implicated in diverse functions in the healthy and diseased brain.

Studies have shown that the supramolecular organization of signaling complexes as nanoclusters regulates signal transduction. Super-resolution microscopy enables us to understand the spatial aspect and microscale features of signaling processes. The use of light (optogenetics) to control and modulate cellular functions has opened up a new arena to explore the cellular functions of biomolecules at a higher spatio-temporal resolution. It enables us to explore the spatio-temporal dynamics of signal transduction. Our study shows optical control of cyclic nucleotide level and the dynamics of downstream effector HCN2 channels at the nanoscale level in a targeted manner. Here, we have developed a new paradigm combining optogenetics with super-resolution microscopy. Optogenetics allows us to modulate the cyclic nucleotide (cAMP) level in a targeted region and super-resolution microscopy permits us to visualize the trafficking and supramolecular organization of molecules at nanoscale resolution. With a flash of light, we modulate the cAMP levels and examine the dynamics of HCN2 channels at both ensemble and single-particle level. We found that optical stimulation to increase intracellular cAMP levels confined the movement of HCN2 channels and segregated them into nanoclusters. We further demonstrated that the alteration in the trafficking and nano-organization of HCN2 channels after binding to cAMP molecules was mediated by their cyclic nucleotide binding domain (CNBD). The integration of two techniques namely, optogenetics and super-resolution microscopy enabled us to modulate and study the biochemical map of signaling complexes at a high spatial-temporal resolution. This paradigm allows altering and comprehending the dynamic nature of nanoscale cellular processes during signal transduction in native and pathophysiological conditions.

A schematic representation illustrating alteration in the cyclic nucleotide level and the molecular arrangement of downstream HCN channels before and after light regulated activation of the protein. Created using Biorender.com

Our study provides a unique way to modulate and investigate the real-time dynamics of signal transduction at the nanoscale level.

What are the possible consequences of these findings for your research area?

Our study provides a unique way to modulate and investigate the real-time dynamics of signal transduction at the nanoscale level. We illustrate how modulations in the level of signaling molecules (cyclic nucleotide) lead to alteration in the nanoscale arrangement and dynamics of downstream molecules. We hope the union of super-resolution microscopy and optogenetics will aid in a better understanding of molecular functions in real-time in native biological and pathophysiological conditions that have not yet been explored at the nanoscale.

 What was the exciting moment (eureka moment) during your research?

In the pursuit of modulating signaling pathways in a spatio-temporal manner and simultaneously visualizing the biochemical map of signaling molecules at the nanoscale level, it was exciting to integrate two different techniques i.e. optogenetics and super-resolution microscopy. The eureka moment was when we controlled the mobility of HCN2 channel in a well-defined region of a live cell via switching on and off blue light. It was exciting to see the synergy between the increase in the level of cyclic nucleotide after illumination and the immobilization of HCN2 molecules to form discrete HCN nanoclusters.

What do you hope to do next?

Since my doctoral study, I have been curious to study precise and coordinated interaction between molecules involved in various signaling cascades. Various pathological and diseased states are associated with the failure or dysregulation of signaling cascades. I would like to use the interface of interdisciplinary research spanning optogenetics, nanobiology, and cell biology to explore the microscale features of signaling complexes in disease conditions via manipulating these cascades in a spatio-temporal manner. I hope, it would enable me to expand the therapeutic targets and genetic approaches to address them.

Where do you seek scientific inspiration?

I have an inclination towards biological sciences and I was always fascinated with the synchronization and coordination between different cells to perform their functions at their respective rhythm inside the human body. Science itself is fascinating, although astonishing research work using advanced technologies to better understand human physiology and for the advancement of human health inspire me to contribute towards the furtherance of healthy human life. Looking up at laureates, my mentors and peers always motivates and encourages me to persistently work smarter and look for interdisciplinary approaches to solve scientific problems. My family is also a big motivational support for me.

How do you intend to help Indian science improve?

I believe science draws everyone’s attention, from kids to elderly. It’s important to have communication between the scientific world and the society to bring awareness and to encourage young generation towards research and development in India. I hope my scientific research is accessible to society and motivates and ignites the curiosity among the next generation. I would be happy to take up new challenges as an independent scientist, share my knowledge, expertise and skills with the next generation, and further contribute to the Indian scientific community.

Reference

Meenakshi Tanwar, Suneel Kateriya, Deepak Nair and  Mini Jose. Optogenetic modulation of real-time nanoscale dynamics of HCN channels using photoactivated adenylyl cyclases. RSC Chem. Biol., 2021, https://doi.org/10.1039/D0CB00124D

Edited by: Ritvi Shah

Leave a Reply

Fill in your details below or click an icon to log in:

WordPress.com Logo

You are commenting using your WordPress.com account. Log Out /  Change )

Google photo

You are commenting using your Google account. Log Out /  Change )

Twitter picture

You are commenting using your Twitter account. Log Out /  Change )

Facebook photo

You are commenting using your Facebook account. Log Out /  Change )

Connecting to %s

%d bloggers like this: