Introduction: Adipose tissue is a complex endocrine organ and active participant in the initiation and progression of metabolic disorders such as obesity and diabetes. While often overlooked, calcium uptake and cycling in the endoplasmic reticulum (ER) are known to play key roles in glucose homeostasis and heat production, a process known as thermogenesis. Despite this knowledge, the regulation of calcium dynamics in adipose and its contribution to disease states remains poorly understood. Here we investigate the function of the micropeptide another regulin (ALN) in modulating the sarcoendoplasmic reticulum ATPase (SERCA2b) pump in subcutaneous adipose tissue and its role in thermogenesis and obesity. Methods: Whole-body reduction of ALN expression in mice was achieved with CRISPR-dCas9-mediated knockdown. Metabolic cohorts (n = 9 per group) were housed at thermoneutrality (30°C) to reduce the contribution of uncoupling protein 1 to thermogenesis. Select mice were also housed at 8°C for 72 hrs to assess cold tolerance. Intraperitoneal glucose and insulin tolerance tests were performed and body composition measured by EchoMRI. Hematoxylin and eosin staining of adipose tissue was used to assess adipocyte size by microscopy. High-resolution respirometry in tissue and isolated mitochondria was measured using an Agilent Seahorse XFe24 Analyzer and Oroboros O2k, respectively. SERCA2b-containing microsomes were isolated via differential centrifugation and heat production measured by isothermal titration calorimetry (ITC; TA instruments). Results: Functionally, genetic knockdown (KD) of ALN increases the uptake of calcium in the ER, while lowering SERCA2b ATPase activity in subcutaneous inguinal white adipose tissue (iWAT), suggesting an altered stoichiometry. In line with these findings, ITC assays demonstrated a decrease in heat production from microsomes lacking ALN. Ing WAT mass relative to whole body weight was increased in the KD cohort, with larger adipocytes as measured by H&E staining. Despite the decreased SERCA ATPase activity, ALN KD mice are cold tolerant, but demonstrate a significant increase in mitochondrial respiration and content at 8°C, possibly indicative of compensatory thermogenic mechanisms in adipose to mitigate the loss of calcium cycling. Discussion: Calcium is the most abundant metal in the human body and, as such, cells must maintain exquisite control of its uptake and compartmentalization to conserve metabolic homeostasis. Bioinformatics and deep sequencing approaches have uncovered novel micropeptides previously misclassified as long noncoding RNAs for which functional roles have not been ascribed. The data presented here demonstrate that ALN is a key regulator of SERCA activity in adipose, the lack of which impairs thermogenesis at the ER as well as insulin tolerance. These findings expand our understanding of the cellular machinery underlying heat production and may provide therapeutic cues for increasing energy expenditure and alleviating metabolic disorders.