Enhanced Autophagy Protects Against ALS Proteotoxicity

By boosting autophagy of misfolded superoxide dismutase 1, researchers protected motor neurons in vitro and in worms from this toxic protein encoded by an ALS gene. A small molecule called SecinH3 speeds up the flow of undesirable proteins through the lysosome, scientists from Children’s Hospital of Philadelphia report in the Jun 17 Journal of Neuroscience. It might prove beneficial in a variety of neurodegenerative diseases where altered proteins accumulate, speculated senior author Robert Kalb.

Neuronal Target.

SecinH3 interferes with the activity of cytohesins, managers of membrane trafficking labeled here in cultured neurons. [Courtesy of Zhai et al., The Journal of Neuroscience, 2015]

Kalb and colleagues were interested in how membrane trafficking participates in ALS pathogenesis. Both the endoplasmic reticulum and lysosomes have been implicated in the disease (see Mar 2009 News story; Sep 2009 News story; Jul 2014 News story). Joint first authors Jinbin Zhai, now at Temple University in Philadelphia, and Lei Zhang investigated the small ADP-ribosylation factors (ARFs) that manage many steps in moving membranes around the cell (reviewed in D’Souza-Schorey and Chavrier, 2006). ARFs hydrolyze GTP; their guanine nucleotide exchange factors, called cytohesins, remove the used-up GDP so ARFs can re-activate quickly. Mammals make at least six ARFs and four cytohesins. When researchers discovered a molecule that inhibits cytohesins (Hafner et al., 2006), the Kalb group was eager to test it out in mSOD1 model systems.

First, they infected cultured rat spinal motor neurons with a viral vector expressing mSOD1. Normally, this kills half the neurons within a week, but SecinH3 kept the cultures alive. The authors also observed neuroprotection when they transduced neurons with dominant-negative cytohesin mutants or interfering RNA against cytohesins.

Next, Zhai and Zhang tried blocking cytohesins in Caenorhabditis elegans expressing the mutant ALS gene. These mSOD1 worms swim slowly, but again, feeding them interfering RNA against cytohesins restored their speed and agility.

The authors suspected the treatments might work by removing misfolded SOD1. In primary neuronal cultures, SecinH3 reduced the concentration of misfolded SOD1, recognized with an antibody specific for the malformed protein, and insoluble, aggregated SOD1.

Based on these results, Zhai and Zhang tested whether SecinH3 treatment enhanced autophagy. They did so by quantifying LC3, an autophagosome component that gets constantly recycled. By stopping autophagy with a compound called Bafilomycin A, the researchers measured how much LC3 is going through the lysosome at a given time. When they treated HeLa or NSC34 cells with SecinH3, they observed more LC3, indicating more traffic on the autophagy pathway.

The researchers still have to work out which of the cell’s many cytohesins and ARFs are crucial to the autophagy process, and how inhibiting them speeds up protein disposal. However, the work could have far-reaching implications. Many other genes involved in ALS, such as profilin 1, also encode proteins that aggregate and might be cleared by cytohesin inactivation, pointed out Daryl Bosco of the University of Massachusetts Medical Center in Worcester, who was not involved in the study. Even in the sporadic form of the disease, some studies have suggested wild-type SOD1 folds improperly (see Oct 2010 News story). “The effect of SecinH3 does not appear specific to mutant SOD1, but rather to the downstream consequences of misfolded proteins,” Bosco said, “I would expect that their findings related to mutant SOD1 in the context of familial ALS are relevant and applicable to aberrantly modified wild type SOD1 in the context of sporadic ALS.”

Beyond ALS, membrane trafficking processes including endocytosis and autophagy are involved in Alzheimer’s, too, said Ralph Nixon of the Nathan Kline Institute in Orangeburg, New York, who was not involved in Kalb’s study. For example, ARF6 mediates the entry of BACE1 into cells (see Aug 2011 News story). “The ability to upregulate autophagy certainly would apply to any disease where a misfolded or abnormal protein would be degraded through the autophagy pathway,” he said. “The big question is, at what level is this upregulation happening?” Lysosomes are impaired in both Alzheimer’s and Parkinson’s, Nixon pointed out, and without knowing how SecinH3 works, researchers cannot be sure if it would be able to fix the problems.

Kalb said scientists have not tried SecinH3 in other models of ALS or neurodegeneration. The compound itself would be no good as a human treatment because it was barely soluble in liquid, he said. Michael Famulok, a study co-author and developer of SecinH3, told Alzforum he mainly intended it as a research tool and has not made much effort to improve its solubility or develop derivatives for human use. The present study illustrates that inhibiting cytohesins in some way might be therapeutic, though, Kalb and colleagues wrote. Other scientists agreed. “They may be on to something very interesting,” commented Robert Brown of the University of Massachusetts Medical school, who did not participate in the study.

To view commentaries, primary articles and linked stories, go to the original posting on Alzforum.org here.

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autophagy disease-als secin topic-preclinical
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