03 Jun 2009

Who will guard the guards? Plato famously grappled with this conundrum, though probably not with this biological version of it: how to protect the proteins in the cell that protect other proteins—the chaperones. That’s a job for modern Greeks. In last month’s PLoS ONE, researchers led by Leonidas Stefanis at the University of Athens Medical School report that chaperone function may be compromised in Parkinson disease. The researchers found that chaperone-driven autophagy (CMA), a fairly specific process for ridding cells of unwanted protein, gets blocked in neurons by mutated or excess α-synuclein. More than that, α-synuclein can trigger a toxic compensation by inducing macroautophagy, a less-targeted form of autophagy. Dysfunctional autophagy has been implicated in Alzheimer (see ARF related news story) and other neurodegenerative diseases (see ARF related news story), and now this work strengthens the link between the Parkinson pathology, as well. The work hints that protecting chaperone-mediated autophagy might be a viable therapeutic approach to the disease. See also yesterday’s story on dementia with Lewy bodies (see ARF related news story).

Stefanis and colleagues previously showed that mutant human (A53T) α-synuclein leads to an accumulation of autophagic vacuoles and that the protein interferes with uptake of CMA substrates in isolated liver lysozomes, but it was not clear if the protein would have the same effect in whole cells, or in neurons for that matter. To test this idea, joint first authors Maria Xilouri and Tereza Vogiatzi expressed α-synuclein constructs in PC12 and SH-SY5Y cells. They found that while the A53T mutant blocked lysosomal degradation of proteins, a mutant lacking the KFERQ peptide motif that targets synuclein for CMA—ΔDQ/A53T ASYN—had no effect. In SH-SY5Y cells that were allowed to differentiate into neurons, the situation was slightly more complex. Wild-type α-synuclein, but not the ΔDQ variant, was capable of blocking CMA-mediated autophagy. A53T synuclein not only blocked CMA, but also the less selective macroautophagy, suggesting a more generalized impairment of protein degradation. Together the results suggest that A53T synuclein blocks CMA in cycling cells and both CMA and macroautophagy in differentiated cells. Wild-type synuclein, which is sufficient to cause Parkinson disease when highly expressed, is only capable of blocking CMA in differentiated cells.

The authors also tested the effects of overexpressing α-synuclein in rat primary cortical neurons. In this scenario, A53T or wild-type synuclein did not block, but instead turned on macroautophagy, but only if the KFERQ peptide motif was present, suggesting that the induction was secondary to an effect on CMA. The proteins were also toxic to the primary neurons, reducing their viability in culture. The ΔDQ variants were less toxic, again suggesting a role for CMA in the toxicity. 3MA, a macroautophagy inhibitor protected the cells, emphasizing the role of macroautophagy in toxicity. The researchers confirmed this by using siRNA to silence expression of ATG5, an essential macroautophagy protein. Knocking down ATG5 dramatically reduced the number of cells killed by A53T α-synuclein. “Taken together, these data indicate that over-expression of A53T α-synuclein induces autophagic death in rat primary cortical cultures that is dependent upon macroautophagy induction, which in turn occurs due to CMA impairment,” write the authors. Interestingly, by inhibiting tyrosine hydroxylase, an enzyme essential for production of dopamine, the authors were able to rescue synuclein-driven macroautophagy in primary cultures. Scientists have puzzled over the selective loss of dopaminergic neurons in Parkinson disease, and this result suggests that dopamine modification of wild-type α-synuclein “may in part be responsible for the reduction of CMA function and increased toxicity,” write the authors.

The authors acknowledge that α-synuclein may exert other toxic effects besides autophagic ones. CMA is activated by α-synuclein monomers, not oligomers or fibrils that may be toxic in their own way. The authors also note that the role of macroautophagy in neurodegeneration remains poorly understood—both lack of and excess autophagy can cause neurodegeneration. But they suggest that CMA might be a potential therapeutic target for PD. “Improving CMA function might not only serve to accelerate α-synuclein degradation, but also to mitigate potential deleterious consequences of aberrant α-synuclein on this system,” they conclude.—Tom Fagan