Open AccessParallel assembly of actin and tropomyosin but not myosin II during de novo actin filament formation in live mice
Author(s) -
Andrius Masedunskas,
Mark A. Appaduray,
Christine A. Lucas,
María Lastra Cagigas,
Marco Heydecker,
Mira Holliday,
Joyce Meiring,
Jeff Hook,
Anthony J. Kee,
Melissa White,
Paul Thomas,
Yingfan Zhang,
Robert S. Adelstein,
Tobias Meckel,
Till Böcking,
Roberto Weigert,
Nicole S. Bryce,
Peter W. Gunning,
Edna C. Hardeman
Publication year - 2018
Publication title -
journal of cell science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.384
H-Index - 278
eISSN - 1477-9137
pISSN - 0021-9533
DOI - 10.1242/jcs.212654
Subject(s) - biology , myosin , tropomyosin , actin remodeling , actin , microbiology and biotechnology , mdia1 , treadmilling , microfilament , protein filament , cytoskeleton , biophysics , actin cytoskeleton , biochemistry , cell
Many actin filaments in animal cells are co-polymers of actin and tropomyosin. For many of these co-polymers, non-muscle myosin II associates to establish a contractile network. However, the temporal relationship of these 3 proteins in the de novo assembly of actin filaments is not known. Intravital subcellular microscopy of secretory granule exocytosis allows the visualisation and quantitation of the formation of an actin scaffold in real time with the added advantage that it occurs in a living mammal, under physiological conditions. We used this model system to investigate the de novo assembly of actin, tropomyosin Tpm3.1 and myosin IIA on secretory granules in mouse salivary glands. Blocking actin polymerization with cytochalasin D revealed that Tpm3.1 assembly is dependent on actin assembly. We used time-lapse imaging to determine the timing of the appearance of the actin filament reporter LifeAct-RFP and of Tpm3.1-mNeonGreen on secretory granules in LifeAct-RFP transgenic, Tpm3.1-mNeonGreen and myosin IIA-GFP knock-in mice. Our findings are consistent with the addition of tropomyosin to actin filaments shortly after the initiation of actin filament nucleation, followed by myosin IIA recruitment.