JIP4 deficiency causes a lysosomal storage disease arising from impaired cystine efflux
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Abstract
Lysosomes break down macromolecules, clear cellular waste and recycle nutrients such as cystine. We describe a novel mechanism whereby JIP4 regulates lysosomal cystine storage by controlling the abundance of cystinosin (CTNS), the transporter responsible for lysosomal cystine efflux. To this end, JIP4, previously characterized as a motor adaptor and kinase signaling scaffold, suppresses TMEM55B-dependent ubiquitylation of CTNS. Loss of JIP4 reduces CTNS protein levels, leading to lysosomal cystine accumulation and lysosomal storage defects that phenocopy loss of CTNS in both human cells and the renal proximal tubules of JIP4 knockout mice. These phenotypes mirror cystinosis, the lysosomal storage disease caused by CTNS loss-of-function. Our findings thus reveal a fundamental process that controls the efflux of lysosomal cystine and has relevance to understanding human disease arising from JIP4 mutations.
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Images of JIP4 KO cells with LAMP1-GFP endogenous tag at 3 different z-planes. Scale bar = 10 µm.
Wow these look crazy! Very prominent phenotype!
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Immunoprecipitation of CTNS-FLAG and the K->A mutant revealed that WT CTNS-FLAG is modified by HA-Ubiquitin, but the K->A mutant is not. Additionally, after normalizing for the reduced abundance of CTNS-FLAG in JIP4 KO cells, CTNS-FLAG was more ubiquitylated in the absence of JIP4
It looks like in figure 3b that the K->A mutations blocking ubiquitination also rescued the levels of CTNS in the JIP4 KO cells, right? The phrasing here is maybe a bit confusing.
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his pulse-chase strategy revealed that CTNS-RUSH initially trafficked similarly in both WT and JIP4 KO cells as it sequentially exhibited an ER-like distribution before biotin+CHX, a Golgi-like distribution after 1h with biotin+CHX, and a predominantly endosome-like punctate distribution after 2h biotin+CHX. However, after 4h, CTNS was abundant on lysosomes in WT cells but difficult to detect in JIP4 KO cells (Fig. 2k-m). This result indicated that, in the absence of JIP4, CTNS is rapidly degraded upon arrival at lysosomes.
Does this mean that WT and JIP4 KO cells had similar levels of initial expression of CTNS? No defects in transcription/translation/etc.? I assume yes, but may be worth explicitly saying, because since JIP4 is somewhat related to the MAPK pathway, my first thought was that this was related to signaling.
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ystine, the oxidized form of the amino acid cysteine that is predominantly found in lysosomes, accumulated in JIP4 KO cells by over 10-fold, and cysteine accumulated by over 5-fold (Fig. 1e-g; Schulman et al., 1969; Gahl, Bashan, et al., 1982; Abu-Remaileh et al., 2017). However, no other metabolites showed comparable changes.
This figure is super striking!
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These findings broaden understanding of lysosomal function, define a new pathway that results in lysosomal storage disease and shed light on how aberrant cystine storage may contribute to human disease arising from JIP4 loss-of-function mutations.
This is a really interesting mechanistic dissection of the underlying cause of CTNS-involved defects that includes the uncovering of a potential novel disease driver and target, JIP4! Very thorough study and lots of useful information!
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