Single molecule imaging of the central dogma reveals myosin-2A gene expression is regulated by contextual translational buffering.
O'Neil WigganTimothy J StasevichPublished in: bioRxiv : the preprint server for biology (2024)
While protein homeostasis is a hallmark of gene regulation, unraveling the hidden regulatory mechanisms that maintain homeostasis is difficult using traditional methods. To confront this problem, we CRISPR engineered a human cell line with multiple tags in the endogenous MYH9 gene, which encodes the essential and ubiquitous myosin-2A cytoskeletal motor. Using these cells, we imaged MYH9 transcription, translation, and mature mRNA and protein in distinct colors, enabling a full dissection of the central dogma. Our data show that MYH9 transcription is upregulated in an SRF-dependent manner in response to cytoskeletal cues and that MYH9 translation can either buffer or match the transcriptional response depending on context. Upon knockdown of actin-depolymerizing proteins like cofilin, translation efficiency drops by a factor of two to buffer strong transcriptional upregulation, likely to help prevent excessive myosin activity. In contrast, following serum stimulation, translation matches the transcriptional response to readily reestablish steady state. Our results identify contextual translational buffering as an important regulatory mechanism driving stable MYH9 expression. They also demonstrate the power and broad applicability of our cell line, which can now be used to accurately quantify central dogma dynamics in response to diverse forms of cellular perturbations.
Keyphrases
- binding protein
- hypertrophic cardiomyopathy
- transcription factor
- gene expression
- single molecule
- left ventricular
- poor prognosis
- dna methylation
- genome wide identification
- induced apoptosis
- magnetic resonance
- endothelial cells
- high resolution
- cell proliferation
- atomic force microscopy
- crispr cas
- genome editing
- computed tomography
- electronic health record
- magnetic resonance imaging
- copy number
- cell cycle arrest
- heart failure
- induced pluripotent stem cells
- oxidative stress
- endoplasmic reticulum stress
- body mass index
- contrast enhanced
- pluripotent stem cells
- amino acid