Abstract
Asna1, also known as TRC40, is implicated in the delivery of tail-anchored (TA) proteins into the endoplasmic reticulum (ER), in vesicle-mediated transport, and in chaperoning unfolded proteins during oxidative stress/ATP depletion. Here, we show that Asna1 inactivation in pancreatic progenitor cells leads to redistribution of the Golgi TA SNARE proteins syntaxin 5 and syntaxin 6, Golgi fragmentation, and accumulation of cytosolic p62+ puncta. Asna1−/− multipotent progenitor cells (MPCs) selectively activate integrated stress response signaling and undergo apoptosis, thereby disrupting endocrine and acinar cell differentiation, resulting in pancreatic agenesis. Rescue experiments implicate the Asna1 ATPase activity and a CXXC di-cysteine motif in ensuring Golgi integrity, syntaxin 5 localization and MPC survival. Ex vivo inhibition of retrograde transport reproduces the perturbed Golgi morphology, and syntaxin 5 and syntaxin 6 expression, whereas modulation of p53 activity, using PFT-α and Nutlin-3, prevents or reproduces apoptosis in Asna1-deficient and wild-type MPCs, respectively. These findings support a role for the Asna1 ATPase activity in ensuring the survival of pancreatic MPCs, possibly by counteracting p53-mediated apoptosis.
Highlights
Deletion of Asna1 in pancreatic progenitor cells leads to severe pancreatic hypoplasia due to apoptosis Asna1 was broadly expressed in the developing pancreatic epithelium from ∼E10.5 and by E13.5 the expression became prominent in the pro-acinar branch tip cells (Fig. S1A)
Here, we show that inactivation of Asna1 in pancreatic progenitor cells leads to selective apoptosis of multipotent progenitor cells, thereby depleting the pancreatic progenitor cell pool and perturbing subsequent growth and differentiation of the developing pancreatic epithelium, which results in severe pancreatic agenesis
The most striking effect of Asna1 deficiency in pancreatic progenitor cells is the severe pancreatic agenesis that appears to be caused by selective apoptosis of multipotent progenitor cells at E13.5
Summary
Known as TRC40, and its yeast homolog GET3 have been implicated in a variety of cellular processes, including proteasome assembly and degradation of ubiquitylated proteins, growth under oxidative or metal stress, membrane trafficking within the secretory pathway, and insulin secretion (Akahane et al, 2013; Auld et al, 2006; Costanzo et al, 2010; Jonikas et al, 2009; Norlin et al, 2016; Schuldiner et al, 2005; Shen et al, 2003). The proposed Asna target TA proteins syntaxin 5 (Stx5) and syntaxin 6 (Stx6) were redistributed from their Golgi compartments both in Asna mutant β-cells and after pharmacological inhibition of retrograde transport using Retro-2 (Norlin et al, 2016; Stechmann et al, 2010). Re-introduction of an ATPase-deficient mutant of Asna failed to restore Golgi integrity and differentiation of pancreatic progenitor cells lacking Asna, suggesting that the ATPasedependent and TA-targeting activities of Asna are required for pancreatic progenitor cell survival
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