Person:

Hagen, Susan

Autophagy related 16-like 1 (ATG16L1) as a genetic risk factor has exposed the critical role of autophagy in Crohn’s disease (CD)1. Homozygosity for the highly prevalent ATG16L1 risk allele, or murine hypomorphic (HM) activity causes Paneth cell dysfunction2,3. As Atg16l1HM mice do not develop spontaneous intestinal inflammation, the mechanism(s) by which ATG16L1 contributes to disease remains obscure. Deletion of the unfolded protein response (UPR) transcription factor X-box binding protein-1 (Xbp1) in intestinal epithelial cells (IECs), whose human orthologue harbors rare inflammatory bowel disease (IBD) risk variants, results in endoplasmic reticulum (ER) stress, Paneth cell impairment and spontaneous enteritis4. Unresolved ER stress is a common feature of IBD epithelium4,5, and several genetic risk factors of CD affect Paneth cells2,4,6-9. Here we show that impairment in either UPR (Xbp1ΔIEC) or autophagy function (Atg16l1ΔIEC or Atg7ΔIEC) in IECs results in each other’s compensatory engagement, and severe spontaneous CD-like transmural ileitis if both mechanisms are compromised. Xbp1ΔIEC mice exhibit autophagosome formation in hypomorphic Paneth cells, which is linked to ER stress via protein kinase RNA-like endoplasmic reticulum kinase (PERK), elongation initiation factor 2α (eIF2α) and activating transcription factor 4 (ATF4). Ileitis is dependent on commensal microbiota and derives from increased IEC death, inositol requiring enzyme 1α (IRE1α)-regulated NFκB activation and tumor necrosis factor signaling which are synergistically increased when autophagy is deficient. ATG16L1 restrains IRE1α activity and augmentation of autophagy in IECs ameliorates ER stress-induced intestinal inflammation and eases NFκB overactivation and IEC death. ER stress, autophagy induction and spontaneous ileitis emerge from Paneth cell-specific deletion of Xbp1. Genetically and environmentally controlled UPR function within Paneth cells may therefore set the threshold for the development of intestinal inflammation upon hypomorphic ATG16L1 function and implicate ileal CD as a specific disorder of Paneth cells.

Objective: Non-shivering thermogenesis in brown adipose tissue (BAT) plays a central role in energy homeostasis. Thioesterase superfamily member 1 (Them1), a BAT-enriched long chain fatty acyl-CoA thioesterase, is upregulated by cold and downregulated by warm ambient temperatures. Them1−/− mice exhibit increased energy expenditure and resistance to diet-induced obesity and diabetes, but the mechanistic contribution of Them1 to the regulation of cold thermogenesis remains unknown. Methods: Them1−/− and Them1+/+ mice were subjected to continuous metabolic monitoring to quantify the effects of ambient temperatures ranging from thermoneutrality (30 °C) to cold (4 °C) on energy expenditure, core body temperature, physical activity and food intake. The effects of Them1 expression on O2 consumption rates, thermogenic gene expression and lipolytic protein activation were determined ex vivo in BAT and in primary brown adipocytes. Results: Them1 suppressed thermogenesis in mice even in the setting of ongoing cold exposure. Without affecting thermogenic gene transcription, Them1 reduced O2 consumption rates in both isolated BAT and primary brown adipocytes. This was attributable to decreased mitochondrial oxidation of endogenous but not exogenous fatty acids. Conclusions: These results show that Them1 may act as a break on uncontrolled heat production and limit the extent of energy expenditure. Pharmacologic inhibition of Them1 could provide a targeted strategy for the management of metabolic disorders via activation of brown fat.

Objectives: Understanding how loci identified by genome wide association studies (GWAS) contribute to pathogenesis requires new mechanistic insights. Variants within CDKAL1 are strongly linked to an increased risk of developing type 2 diabetes and obesity. Investigations in mouse models have focused on the function of Cdkal1 as a tRNALys modifier and downstream effects of Cdkal1 loss on pro-insulin translational fidelity in pancreatic β−cells. However, Cdkal1 is broadly expressed in other metabolically relevant tissues, including adipose tissue. In addition, the Cdkal1 homolog Cdk5rap1 regulates mitochondrial protein translation and mitochondrial function in skeletal muscle. We tested whether adipocyte-specific Cdkal1 deletion alters systemic glucose homeostasis or adipose mitochondrial function independently of its effects on pro-insulin translation and insulin secretion. Methods: We measured mRNA levels of type 2 diabetes GWAS genes, including Cdkal1, in adipose tissue from lean and obese mice. We then established a mouse model with adipocyte-specific Cdkal1 deletion. We examined the effects of adipose Cdkal1 deletion using indirect calorimetry on mice during a cold temperature challenge, as well as by measuring cellular and mitochondrial respiration in vitro. We also examined brown adipose tissue (BAT) mitochondrial morphology by electron microscopy. Utilizing co-immunoprecipitation followed by mass spectrometry, we performed interaction mapping to identify new CDKAL1 binding partners. Furthermore, we tested whether Cdkal1 loss in adipose tissue affects total protein levels or accurate Lys incorporation by tRNALys using quantitative mass spectrometry. Results: We found that Cdkal1 mRNA levels are reduced in adipose tissue of obese mice. Using adipose-specific Cdkal1 KO mice (A-KO), we demonstrated that mitochondrial function is impaired in primary differentiated brown adipocytes and in isolated mitochondria from A-KO brown adipose tissue. A-KO mice displayed decreased energy expenditure during 4 °C cold challenge. Furthermore, mitochondrial morphology was highly abnormal in A-KO BAT. Surprisingly, we found that lysine codon representation was unchanged in Cdkal1 A-KO adipose tissue. We identified novel protein interactors of CDKAL1, including SLC25A4/ANT1, an inner mitochondrial membrane ADP/ATP translocator. ANT proteins can account for the UCP1-independent basal proton leak in BAT mitochondria. Cdkal1 A-KO mice had increased ANT1 protein levels in their white adipose tissue. Conclusions: Cdkal1 is necessary for normal mitochondrial morphology and function in adipose tissue. These results suggest that the type 2 diabetes susceptibility gene CDKAL1 has novel functions in regulating mitochondrial activity.