Panoply™ Human SYK Knockdown Stable Cell Line

Spleen tyrosine kinase (SYK) plays a crucial role in inflammatory and adaptive immune responses, and is therefore likely associated with neuroinflammation observed in various neurodegenerative diseases. SYK also regulates the production of β-amyloid (Aβ) and the hyperphosphorylation of Tau protein, which is associated with these diseases. Furthermore, in Alzheimer's disease (AD) patients and AD mouse models, SYK overactivation occurs in partially activated microglia, dystrophic neurites surrounding Aβ deposits, and neurons affected by Tau pathology. SYK activation increases Tau phosphorylation and accumulation, suggesting that SYK may be a potential therapeutic target for AD. Here, researchers found that SYK inhibition increases autophagic Tau protein degradation without affecting Tau protein production. Using neuron-like SH-SY5Y cells, researchers demonstrated that SYK is located upstream of the mammalian target of rapamycin (mTOR) pathway, and that pharmacological inhibition or knockdown of SYK reduces mTOR pathway activation and increases autophagic Tau protein degradation. Interestingly, in a mouse model of tau protein disease, long-term inhibition of SYK significantly reduced tau protein accumulation, neuroinflammation, neuronal and synaptic loss, and reversed autophagy defects. These results further indicate that SYK upregulation observed in the brains of Alzheimer's patients leads to autophagy clearance defects, which in turn result in the accumulation of pathogenic tau protein.

To validate the data obtained from SYK pharmacological inhibition using BAY61 and to ensure that the observed effects were mediated by SYK inhibition, researchers constructed SYK-knockdown SH-SY5Y cells. Compared to control cells, the total SYK (t-SYK) level was significantly reduced in SYK-knockdown SH-SY5Y cells (Figure 1). Researchers found that SYK knockdown significantly reduced the levels of p-mTOR (Ser-2448), p-S6K (Thr-389), p-Akt (Ser-473), p-Tau (Ser-396/404), and t-Tau (Figures 1A and B), mimicking the effect of the SYK inhibitor BAY61 in SH-SY5Y cells. Interestingly, although the ratio of LC3-I to LC3-II remained unchanged, both were significantly reduced in SYK-knockdown cells (Figure 1A). These data suggest that gene inhibition of SYK expression leads to increased autophagy initiation (conversion of LC3-I to LC3-II) and enhanced lysosomal LC3-II degradation. To further validate this hypothesis, researchers treated control and SYK knockdown cells with 200 μM of the lysosomal inhibitor CQ for 24 hours. In the presence of CQ, the LC3-II/I ratio in the SYK knockdown group was significantly increased, similar to the results observed in SH-SY5Y cells treated with CQ and BAY61, indicating increased autophagy flux and suggesting that SYK may influence autophagy initiation and lysosomal degradation (Figure 1C).

Figure 1. SYK knockdown mimics pharmacological inhibition of SYK and decreases total Tau levels. (Schweig J E, et al., 2019)