Rapid Homeostatic Turnover of Embryonic ECM during Tissue Morphogenesis

Abstract

Highlights d Labeled ECM in fly embryos can be examined from initiation to homeostasis d Quantifying ECM levels to homeostasis allows for modeling of basal turnover rate d Embryonic ECM has a half-life of 10 h, which was confirmed by pulse-chase analysis d Inhibiting MMPs or ECM interactions alters the basal turnover rate

Mechano-chemical enforcement of tendon apical ECM into nano-filaments during Drosophila flight muscle development

“…Recent studies have uncovered ECM modulation mechanisms by regulated synthesis and secretion, subunit assembly, chemical modification, and proteolytic degradation. [60][61][62][63] Qsm is a new class of molecules regulating apical ECM dynamics by targeting a subset of ZPD proteins and is expected to provide a new direction in the developmental control of ECM dynamics.…”

Protocol for intervention-free quantification of protein turnover rate by steady-state modeling

“…Two datasets are used as examples in this protocol, related to Collagen IV (ColIV) and Nidogen (Ndg) expression in the Drosophila embryo (see Material S3 ). mRNA levels were quantified from an RNA-seq time course ( Graveley et al., 2011 ) and protein levels were obtained by measuring the fluorescence intensity of GFP protein-trap lines during development ( Matsubayashi et al., 2020 ) (see the data acquisition section for more details). It should be noted that this approach is not limited to in vivo models and to the experimental methods used here.…”

“…The model can be mathematically described as follows: where P(t) is the protein expression over time t, M(t) is the mRNA profile over time, S p is the constant rate of protein synthesis, and D p the constant rate of protein degradation ( Matsubayashi et al., 2020 ; Tchourine et al., 2014 ). This corresponds to hypothesizing that the net change in protein levels over time is determined by synthesis minus degradation, with the amount of synthesis and degradation proportional to the mRNA levels and the protein levels, respectively.…”

“…This model was successfully employed to quantify turnover during Drosophila embryogenesis, and we hypothesize that it will be applicable to diverse in vivo or in vitro systems. For complete details on the use and execution of this protocol, please refer to Matsubayashi et al. (2020) .…”

“…For complete details on the use and execution of this protocol, please refer to Matsubayashi et al. (2020) .…”