Abstract
Phosphatase and tensin homolog (Pten) catalyzes the reverse reaction of PI3K by dephosphorylating PIP3 to PIP2. This negatively regulates downstream Akt/mTOR/S6 signaling resulting in decreased cellular growth and proliferation. Co-injection of a lentivirus knocking Pten down with a control lentivirus allows us to compare the effects of Pten knockdown between individual neurons within the same animal. We find that knockdown of Pten results in neuronal hypertrophy by 21 days post-injection. This neuronal hypertrophy is correlated with increased p-S6 and p-mTOR in individual neurons. We used this system to test whether an environmental factor that has been implicated in cellular hypertrophy could influence the severity of the Pten knockdown-induced hypertrophy. Implantation of mini-osmotic pumps delivering fatty acids results in increased neuronal hypertrophy and p-S6/p-mTOR staining. These hypertrophic effects were reversed in response to rapamycin treatment. However, we did not observe a similar increase in hypertrophy in response to dietary manipulations of fatty acids. Thus, we conclude that by driving growth signaling with fatty acids and knocking down a critical regulator of growth, Pten, we are able to observe an additive morphological phenotype of increased soma size mediated by the mTOR pathway.
Highlights
Phosphatase and tensin homolog on chromosome 10 (Pten), a phosphatase catalyzing the reverse reaction of phosphatidylinositol 3-kinases (PI3K), cleaves phosphate from phosphatidylinositol (3,4,5)-triphosphate to yield phosphatidylinositol (4,5)-bisphosphate
LENTIVIRAL Pten KNOCKDOWN RESULTS IN NEURONAL HYPERTROPHY C57BL/6 mice at 6–8 weeks of age were co-injected with an FUGW-based lentivirus expressing both GFP and an shRNA targeting the Pten coding region (GFP shPten) and a control virus www.frontiersin.org expressing only mCherry (Lois et al, 2002; Luikart et al, 2011b)
Since we found increased p-S6 and p-mTOR signaling in Pten knockdown neurons, we hypothesized that rapamycin, a potent mTOR inhibitor, would reverse or inhibit this neuronal hypertrophy
Summary
Phosphatase and tensin homolog on chromosome 10 (Pten), a phosphatase catalyzing the reverse reaction of phosphatidylinositol 3-kinases (PI3K), cleaves phosphate from phosphatidylinositol (3,4,5)-triphosphate to yield phosphatidylinositol (4,5)-bisphosphate. Pten inhibits signaling through the Akt/mTOR/S6 pathway downstream of growth factor receptors (Laplante and Sabatini, 2012). The neuronal hypertrophy seen in mouse models parallels the macrocephaly that characterizes a subset of patients with Pten mutations and Bannayan-RileyRuvalcaba syndrome, Cowden Syndrome, and Autism Spectrum Disorder (ASD) (Longy et al, 1998; Goffin et al, 2001; Butler et al, 2005; McBride et al, 2010). A number of other proteins related to growth factor signaling including Neurofibromin 1 (NF1), Tuberous Sclerosis Complex 1 and 2 (TSC1/2), and fragile X mental retardation protein (FMRP) are associated with ASD and other co-morbidities such as epilepsy, mental retardation, and tumor formation (Kelleher and Bear, 2008; Bourgeron, 2009). Multifactorial genetic and environmental factors likely contribute to the etiology of ASD (Persico and Bourgeron, 2006), little is known about how gene/gene and gene/environment interactions influence the expression of cellular phenotypes
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