Fate-mapping analysis using Rorb-IRES-Cre reveals apical-to-basal gradient of Rorb expression in mouse cochlea

Area of Science:

• Genetics
• Neuroscience
• Otolaryngology

Background:

• Conditional loss-of-function studies are crucial for understanding gene function, especially to avoid embryonic lethality.
• The Cre/Loxp system is widely used, but a lack of inner ear-specific Cre strains hinders research.
• Existing methods often result in lethality, complicating conditional knockout studies in the auditory system.

Purpose of the Study:

• To develop and characterize a novel Rorb-IRES-Cre knockin mouse model for inner ear research.
• To assess the Cre recombinase activity driven by the Rorb promoter in the developing cochlea and vestibule.
• To enable precise spatial and temporal control of gene manipulation within the inner ear.

Main Methods:

• Generation of a Rorb-IRES-Cre knockin mouse line.
• Crossing with Rosa26-CAG-LSL-tdTomato reporter mice to visualize Cre activity.
• Analysis of tdTomato expression patterns in cochlear and vestibular tissues at various embryonic and postnatal developmental stages (E12.5 to P30).
• Confirmation of Rorb mRNA and protein expression in auditory cells.

Main Results:

• Cre-mediated recombination, indicated by tdTomato expression, was observed in the cochlea with an apical-to-basal gradient.
• Expression was initially restricted to the apical turn and expanded to middle and basal turns over time.
• Auditory hair cells and supporting cells in all cochlear turns showed significant recombination by P15 and P30.
• Vestibular cells showed minimal recombination, appearing only at later postnatal stages (P15-P30).
• Rorb mRNA and protein were confirmed in cochlear hair cells and supporting cells at P30.

Conclusions:

• The Rorb-IRES-Cre mouse line effectively recapitulates Rorb expression patterns in the inner ear.
• Rorb Cre activity exhibits a distinct apical-to-basal gradient within the cochlea.
• This model allows for the discrimination of gene functions between cochlear and vestibular regions.
• It also facilitates the study of gene function in low-frequency (apical) versus high-frequency (basal) regions of the cochlea.