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Embryonic lethality and defective mammary gland development of activator-function impaired conditional knock-in Erbb3
Kate Senger1, Wenlin Yuan1, Meredith Sagolla2
1Department of Molecular Biology Genentech South San Francisco California USA.
Abstract:
ERBB3 is a pseudokinase domain-containing member of the ERBB family of receptor tyrosine kinases (RTKs). Following ligand binding, ERBB receptors homo- or hetero-dimerize, leading to a head-to-tail arrangement of the intracellular kinase domains, where the "receiver" kinase domain of one ERBB is activated by the "activator" domain of the other ERBB in the dimer. In ERBB3, a conserved valine at codon 943 (V943) in the kinase C-terminal domain has been shown to be important for its function as an "activator" kinase in vitro. Here we report a knock-in mouse model where we have modified the endogenous Erbb3 allele to allow for tissue-specific conditional expression of Erbb3 (Erbb3 ). Additionally, we generated an Erbb3 (Erbb3 ) conditional knock-in mouse model where the conserved aspartate in the DFG motif of the pseudokinase domain was mutated to abolish any potential residual kinase activity. While Erbb3 animals developed normally, homozygous Erbb3 expression during development resulted in embryonic lethality. Further, tissue specific expression of Erbb3 in the mammary gland epithelium following its activation using MMTV-Cre resulted in delayed elongation of the ductal network during puberty. Single-cell RNA-seq analysis of Erbb3 mammary glands showed a reduction in a specific subset of fibrinogen-producing luminal epithelial cells.
Insights
Researchers created knock-in mice to study ERBB3, a pseudokinase. Homozygous ERBB3 mutations caused embryonic lethality and affected mammary gland development, revealing ERBB3
Area of Science:
- Molecular Biology
- Developmental Biology
- Genetics
Background:
- ERBB3 is a pseudokinase within the ERBB family of receptor tyrosine kinases (RTKs).
- ERBB receptor activation involves dimerization and trans-activation of intracellular kinase domains.
- A specific valine (V943) in ERBB3 is crucial for its activator function in vitro.
Purpose of the Study:
- To generate and characterize conditional knock-in mouse models for ERBB3.
- To investigate the in vivo function of ERBB3, particularly its role in development and tissue-specific contexts.
- To elucidate the consequences of impaired ERBB3 kinase activity and expression.
Main Methods:
- Generation of conditional knock-in mice with modified Erbb3 alleles.
- Creation of a pseudokinase-dead Erbb3 mutant (Erbb3-dn) by mutating the DFG motif.
- Utilizing MMTV-Cre for tissue-specific conditional expression in mammary glands.
- Employing single-cell RNA-sequencing (scRNA-seq) for molecular profiling.
Main Results:
- Erbb3(V943) knock-in animals developed normally.
- Homozygous Erbb3(dn) expression led to embryonic lethality.
- Conditional Erbb3(dn) expression in mammary glands impaired ductal network elongation during puberty.
- scRNA-seq revealed a reduction in specific fibrinogen-producing luminal epithelial cells in Erbb3(dn) mammary glands.
Conclusions:
- ERBB3 kinase activity is essential for embryonic development.
- ERBB3 plays a critical role in mammary gland morphogenesis, influencing ductal network formation.
- Loss of ERBB3 function impacts specific epithelial cell populations within the mammary gland.
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