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A Ser75-to-Asp phospho-mimicking mutation in Src accelerates ageing-related loss of retinal ganglion cells in mice
Kenji Kashiwagi1, Sadahiro Ito2,3, Shuichiro Maeda2
1Department of Ophthalmology, University of Yamanashi, Yamanashi, 409-3898, Japan.
Abstract:
Src knockout mice show no detectable abnormalities in central nervous system (CNS) post-mitotic neurons, likely reflecting functional compensation by other Src family kinases. Cdk1- or Cdk5-dependent Ser75 phosphorylation in the amino-terminal Unique domain of Src, which shares no homology with other Src family kinases, regulates the stability of active Src. To clarify the roles of Src Ser75 phosphorylation in CNS neurons, we established two types of mutant mice with mutations in Src: phospho-mimicking Ser75Asp (SD) and non-phosphorylatable Ser75Ala (SA). In ageing SD/SD mice, retinal ganglion cell (RGC) number in whole retinas was significantly lower than that in young SD/SD mice in the absence of inflammation and elevated intraocular pressure, resembling the pathogenesis of progressive optic neuropathy. By contrast, SA/SA mice and wild-type (WT) mice exhibited no age-related RGC loss. The age-related retinal RGC number reduction was greater in the peripheral rather than the mid-peripheral region of the retina in SD/SD mice. Furthermore, Rho-associated kinase activity in whole retinas of ageing SD/SD mice was significantly higher than that in young SD/SD mice. These results suggest that Src regulates RGC survival during ageing in a manner that depends on Ser75 phosphorylation.
Insights
Phosphorylation of Src protein at Ser75 is crucial for retinal ganglion cell (RGC) survival in aging mice. Mutants mimicking this phosphorylation show RGC loss, unlike non-phosphorylatable or wild-type mice, indicating Src
Area of Science:
- Neuroscience
- Cell Biology
- Ophthalmology
Background:
- Src knockout mice show no central nervous system (CNS) abnormalities, suggesting functional compensation by other Src family kinases.
- Serine 75 (Ser75) phosphorylation in the Src unique domain regulates active Src stability and is dependent on Cdk1 or Cdk5.
- The specific role of Src Ser75 phosphorylation in CNS neurons, particularly in retinal ganglion cells (RGCs), remains unclear.
Purpose of the Study:
- To investigate the role of Src Ser75 phosphorylation in the survival of CNS neurons, specifically RGCs, during aging.
- To determine if specific phosphorylation states of Src at Ser75 affect age-related RGC loss.
- To explore the downstream mechanisms, such as Rho-associated kinase activity, involved in Src-mediated RGC regulation.
Main Methods:
- Generation of two mutant mouse lines: Ser75Asp (SD), a phospho-mimicking mutant, and Ser75Ala (SA), a non-phosphorylatable mutant.
- Comparative analysis of RGC numbers in young and aging SD/SD, SA/SA, and wild-type (WT) mice.
- Assessment of inflammation and intraocular pressure to rule out confounding factors in RGC loss.
- Measurement of Rho-associated kinase activity in retinal tissues.
Main Results:
- Aging SD/SD mice exhibited significant RGC loss compared to young SD/SD mice, without inflammation or elevated intraocular pressure.
- SA/SA and WT mice showed no age-related RGC loss, indicating that non-phosphorylatable Src or normal Src levels protect against RGC degeneration.
- RGC loss in SD/SD mice was more pronounced in peripheral than mid-peripheral retinal regions.
- Rho-associated kinase activity was significantly elevated in aging SD/SD mice retinas.
Conclusions:
- Src protein regulates RGC survival during aging.
- Ser75 phosphorylation of Src is critical for this regulatory function, as mimicking phosphorylation leads to RGC loss.
- Age-related RGC loss in the phospho-mimicking mutant is associated with increased Rho-associated kinase activity, suggesting a potential mechanism.
