Faculty Profile

Mark S. Shapiro, Ph.D.

Research Interests

What are ion channels? - The nervous system is a vast array of connected cells, called neurons, which are responsible for organizing and directing the activities of all animals. The heart is composed of cardiomyocytes that are electrically connected to control and sustain the cardiovascular system. Smooth muscle cells control the tone of blood vessels and airways. These neurons, cardiomyocytes and muscle cells work by using a combination of electrical and chemical signals. The electrical signals use the charged salt atoms (ions) of body fluids that are our heritage of evolving from the sea. The movement of ions in and out of cells is mediated by ion channels, macromolecular proteins that span the lipid membranes of cells, and selectively permit the passage of particular ions. The ionic currents carried by ion channels underlie the basic electrical signals that allow the nervous and cardiovascular systems to function. We are interested in the physiology, regulation and functional role of ion channels.

Many signaling pathways modulate ion channels.Since ion channels are so critical to the function of excitable cells, a vast system of neurotransmitters and hormones regulate the physiology of excitable cells by acting on specific receptors in the cell membrane. The intracellular actions of these receptors are often mediated by G proteins, biological molecular switches that transduce the extracellular presence of a neurotransmitter into the modulation of some intracellular target. In neurons and cardiomyocytes, and smooth muscle cells, as well as many other types of cells, these G protein-mediated signaling pathways frequently act on ion channels. Our laboratory focuses on these G-protein signaling pathways that act on two important types of ion channels. The first makes an important neuronal K+ current called the M current, called such because it is strongly modulated by muscarinic acetylcholine receptor agonists. The genes underlying M currents have been identified, and several different subunits of the KCNQ (Kv7) K+ channel family, consisting of KCNQ1-5 (Kv7.1-7.5), come together to form M-type K+ channels. The second are voltage-gated Ca2+ channels, whose currents drive exocytosis and the release of neurotransmitter at nerve terminals. Ca2+ channels are composed of pore-forming α-subunits, along with auxiliary ß, α2δ and ɣ subunits. By modulating Ca2+ channel activity, synaptic transmission is thus directly regulated. Modulation of the M-type K+ currents plays a strong role in regulating the excitability of neurons, the firing of action potentials and the tone of smooth muscle. Many different receptors act via G proteins to modulate the M current and Ca2+ currents. We know that these G-protein pathways use intracellular second messengers, and that the intracellular mechanisms they use are often distinct. One goal of the lab is to elucidate the mechanisms of this receptor-specificity of action.
Shapiro research

Model accounting for receptor-specific phosphoinositide and Ca2+ signals in sympathetic neurons. All receptors activate PLCß, which in turn hydrolyses PIP2 to IP3 and diacylglycerol (DAG). The released Ca2+ is regulated by the IP3R binding protein, IRBIT, which sets a "threshold" for [IP3] sufficient to open IP3Rs. Stimulation of M1 muscarinic acetylcholine receptors (left) is ineffective in producing cytoplasmic Ca2+signals since the IP3 produced is too far away from IP3Rs; thus, [IP3] at the IP3R is too low to overcome the IRBIT threshold. Bradykinin B2 and purinergic P2Y receptors (right) produce robust cytoplasmic Ca2+ signals due to their spatial co-localization with IP3Rs where [IP3] is sufficiently high. Via neuronal Ca2+ sensor-1 (NCS-1), bradykinin and purinergic, but not muscarinic, stimulation accelerates PI 4IIIß-kinase activity. Via DAG-kinase conversion to phosphatidic acid (PA), the produced DAG increases PI(4)P 5-kinase (PI4P-K) activity, in concert with Rho-family proteins and Rho-kinase (R-K). PI(4)P 5-kinase activity is also increased by bradykinin and purinergic stimulation, but for clarity this is only shown for muscarinic. Acceleration of both PI 4IIIß- and PI(4)P 5-kinases is required to increase PIP2 synthesis that compensates for consumption of PIP2 by PLC.

We study these channels using preparations of primary sympathetic and sensory neurons, cardiomyocytes and smooth muscle cells, and using heterologous mammalian expression systems. Our major approaches include patch clamp electrophysiology, along with techniques of molecular biology and biochemistry, to probe the molecular mechanism of ion channel modulation. We also perform single-cell living imaging experiments, using confocal and TIRF microscopy, specialized lasers and highly-sensitive cameras. Such techniques include Ca2+ imaging, Förster Resonance Energy Transfer (FRET), in vivo brain imaging and GFP-tagged reporter fluorescent probes, to directly observe the creation, reduction or translocation in individual cells of many putative second messengers.

Recently, we have begun to study the clustering of signaling complexes by the scaffolding protein, A-kinase anchoring protein (AKAP)79/150. AKAPs organize "signalosomes" that include kinases, phosphatases, calmodulin, phosphoinositides, receptors and ion channels. Using transgenic "knock-out" mice, patch-clamp electrophysiology, FRET and GFP translocation experiments, we have shown AKAP79/150 to orchestrate both "short-term" modulation of M-type K+ and L-type Ca2+ channels in a receptor-specific manner, as well as "longer-term" regulation of KCNQ2/3 gene transcription, via NFATc1-4 transcription factors, acting as a negative-feedback loop that prevents hyperexcitabilty in the brain, such as seen during epilepsy or chronic pain. An example experiment is shown below.

Shapiro research

With the combined use of biophysics, molecular biology, biochemistry and single-cell imaging, this lab works toward the identification of the relevant signaling molecules in modulation of ion channels and the understanding of the precise mechanisms they use.


Lab Team

  • Crystal Archer
    Crystal Archer
  • Chase Carver
  • MaryAnn Hobbs
  • Isamar Sanchez
  • Rafael Veraza
  • Fabio Vigil
Education B.A., Physics, University of Chicago, 1984 Ph.D., Physiology, Rush University Medical Center, 1991
Selected Publications

Zhang, J., Carver, C.C., Choveau F., and M.S. Shapiro.  (2016). Clustering and functional coupling of diverse ion channels and signaling proteins revealed by super-resolution STORM microscopy in neurons.  Neuron.  92(2):461-478.

Choveau, F., Bierbower, S.M. and M. S. Shapiro.  (2016). The role of the carboxyl terminus helix C-D linker in regulating KCNQ3 K+ current amplitudes by controlling channel trafficking.  PloS One, 10(12):Dec. 21.

Zhang, J. and M.S. Shapiro.  (2015). Novel roles of AKAP79/150 in orchestrating multi-protein signaling complexes in brain and peripheral nerve.  Journal of Physiology 594(1):31-37.