Mining the molecular noise: fluorescence fluctuation image analysis of microscopy and super-resolution microscopy images reveals biomolecule interactions & transport maps in living cells

Speaker

Abstract

The transport properties of biomolecules in cells can reveal a great deal about the functional interactions regulating cells at the molecular level. Various biophysical methods have been developed to measure these properties in cells, although most have relied on fluorescence microscopy imaging as the window for measurement of labeled macromolecules in living cells. Image correlation methods are an extension of fluorescence fluctuation spectroscopy that can measure protein-protein interactions and macromolecular transport properties from input fluorescence microscopy images of living cells. These approaches are based on space and time correlation analysis of fluctuations in fluorescence intensity within images recorded as a time series using a fluorescence or super-resolution microscope. I will introduce spatio-temporal image correlation spectroscopy (STICS) and its 2 color cross-correlation variant (STICCS) and show how the analysis can reveal hidden coupling between retrograde cellular actin flows and the plasma membrane lipids for activated Jurkat cells. I will then describe the application of the STICS and pair vector correlation for measuring cellular waves of adhesion related macromolecules talin and vinculin as well as cytoskeletal actin between assembling and disassembling podosomes in dendritic immune cells. Podosomes are cylindrical membrane complexes with an integrin adhesive ring and an actin rich core that are associated with cellular migration and invasion in specific cell types. EM and super-resolution microscopy of cells shows radial actin filaments that connect neighboring podosomes. The image correlation analysis combined with pharmacological perturbation experiments show that podosome turnover is coordinated within local clusters in cells with a correlation length scale extending to next nearest neighbor podosomes. Moreover, recent work pairing the analysis with live cell super-resolution microscopy reveals that the dynamic coordination between podosomes differs for cells on soft versus stiff substrates which provides clues to the mechanistic function of podosomes.


Show Resources
You May Also Like
SEP 14, 2021 7:00 AM PDT
C.E. CREDITS
SEP 14, 2021 7:00 AM PDT
Date: September 14, 2021 Time: 7am PDT, 10am EDT, 4pm CEST A conventional thermal cycler has long been a commodity product in the lab and end-point PCR techniques can be completed almost wit...
SEP 17, 2021 12:00 PM CST
C.E. CREDITS
SEP 17, 2021 12:00 PM CST
Date: September 16, 2021 Time: 9:00pm (PDT), 12:00am (EDT) 3D cellular models like organoids and spheroids offer an opportunity to better understand complex biology in a physiologically rele...
DEC 01, 2021 7:00 AM PST
C.E. CREDITS
DEC 01, 2021 7:00 AM PST
Date: December 01, 2021 Time: 7:00am (PST), 10:00am (EST) In the era of immuno-oncology, there is a growing need for the identification of new biomarkers predictive for sensitivity to anti-P...
OCT 12, 2021 9:00 AM PDT
C.E. CREDITS
OCT 12, 2021 9:00 AM PDT
Date: October 12, 2021 Time: 9:00am (PDT), 12:00pm (EDT) SCIEX’s next-generation Biologics Explorer software is an innovative platform for the comprehensive and deep characterization o...
NOV 09, 2021 11:00 AM PST
C.E. CREDITS
NOV 09, 2021 11:00 AM PST
Date: November 09, 2021 Time: 11:00am (PDT), 02:00pm (EDT) Clinical translation of human pluripotent stem cells (hPSCs) requires advanced strategies that ensure safe and robust long-term gro...
JUN 24, 2021 10:00 AM PDT
C.E. CREDITS
JUN 24, 2021 10:00 AM PDT
Date: June 24, 2021 Time: 10:00am PDT The Chan Zuckerberg Initiative (CZI) was founded to help solve some of society’s toughest challenges— from eradicating disease and improving...
Loading Comments...
Show Resources