Illuminating the Cell’s Biochemical Activity Architecture

Abstract

All cellular behaviors arise through the coordinated actions of numerous intracellular biochemical pathways. Over the past twenty years, efforts to probe intracellular biochemical processes have undergone a fundamental transformation brought about by advances in fluorescence imaging, such as the development of genetically encoded fluorescent reporters and new imaging technologies; the impact of these approaches on our understanding of the molecular underpinnings of biological function cannot be understated. In particular, the ability to obtain information on the spatiotemporal regulation of biochemical processes unfolding in real time in the native context of a living cell has crystallized the view, long a matter of speculation, that cells achieve specific biological outcomes through the imposition of spatial control over the distribution of various biomolecules, and their associated biochemical activities, within the cellular environment. Indeed, the compartmentalization of biochemical activities by cells is now known to be pervasive and to span a multitude of spatial scales, from the length of a cell to just a few enzymes. In this Perspective, part of this special issue on “Seeing into cells”, we highlight several recent imaging studies that provide detailed insights into not just where molecules are but where molecules are active within cells, offering a glimpse into the emerging view of biochemical activity architecture as a complement to the physical architecture of a cell.

Spatial regulation of AMPK signaling revealed by a sensitive kinase activity reporter

“…Genetically encoded reporters are powerful tools for interrogating the precise spatiotemporal regulation of signaling pathways 12 , 13 . Addition of localization tags for cellular organelles and compartments to kinase activity reporters enables subcellular resolution of kinase activities in single cells, allowing for the elucidation of compartment-specific signaling mechanisms 14 , 15 .…”

Spatial regulation of AMPK signaling revealed by a sensitive kinase activity reporter

“…Mechanistically, the importance of upstream kinases for ADAM site activation remains controversial 48,49 , as ADAM site activators protect AMPK from dephosphorylation by phosphatases 3,48,50 . As kinase and phosphatase activities can vary based on location 12,13 , we hypothesized there could be spatially distinct differences in allosteric activator induced AMPK activity. To test this hypothesis, we set out to use ExRai AMPKAR to profile spatiotemporal AMPK activity in response to allosteric activation via the ADAM site and determine the importance of the upstream kinase LKB1 for spatially defined, allosterically activated AMPK.…”

“…Genetically encoded reporters are powerful tools for interrogating the precise spatiotemporal regulation of signaling pathways 12,13 . Addition of localization tags for cellular organelles and compartments to kinase activity reporters enables subcellular spatial resolution of kinase activities in single cells, allowing for the elucidation of compartment-specific signaling mechanisms 14,15 .…”

Introducing the “Seeing into Cells” Special Issue

“…Mikhail Shapiro’s contribution (Caltech) takes this special issue beyond imaging to include control of cellular function using acoustic and magnetic methods . Jin Zhang (UC San Diego) focuses on recent studies to identify where molecules are active within cells, efforts that provide much clearer glimpses into cellular architectureas featured on this special issue’s cover . Finally, Dirk Trauner (New York University) addresses the difficulties in targeting receptors for drug development by covering the use of tethered photopharmacology …”