MAF1 is a global repressor of RNA polymerase III transcription that regulates the expression of highly abundant noncoding RNAs in response to nutrient availability and cellular stress. Thus, MAF1 function is thought to be important for metabolic economy. Here we show that a whole-body knockout of Maf1 in mice confers resistance to diet-induced obesity and nonalcoholic fatty liver disease by reducing food intake and increasing metabolic inefficiency. Energy expenditure in Maf1 −/− mice is increased by several mechanisms. Precursor tRNA synthesis was increased in multiple tissues without significant effects on mature tRNA levels, implying increased turnover in a futile tRNA cycle. Elevated futile cycling of hepatic lipids was also observed. Metabolite profiling of the liver and skeletal muscle revealed elevated levels of many amino acids and spermidine, which links the induction of autophagy in Maf1 −/− mice with their extended life span. The increase in spermidine was accompanied by reduced levels of nicotinamide N-methyltransferase, which promotes polyamine synthesis, enables nicotinamide salvage to regenerate NAD + , and is associated with obesity resistance. Consistent with this, NAD + levels were increased in muscle. The importance of MAF1 for metabolic economy reveals the potential for MAF1 modulators to protect against obesity and its harmful consequences.

Human-specific duplications at chromosome 16p11.2 mediate recurrent pathogenic 600 kbp BP4-BP5 copy-number variations, which are among the most common genetic causes of autism. These copy-number polymorphic duplications are under positive selection and include three to eight copies of BOLA2, a gene involved in the maturation of cytosolic iron-sulfur proteins. To investigate the potential advantage provided by the rapid expansion of BOLA2, we assessed hematological traits and anemia prevalence in 379,385 controls and individuals who have lost or gained copies of BOLA2: 89 chromosome 16p11.2 BP4-BP5 deletion carriers and 56 reciprocal duplication carriers in the UK Biobank. We found that the 16p11.2 deletion is associated with anemia (18/89 carriers, 20%, p ¼ 4eÀ7, OR ¼ 5), particularly iron-deficiency anemia. We observed similar enrichments in two clinical 16p11.2 deletion cohorts, which included 6/63 (10%) and 7/20 (35%) unrelated individuals with anemia, microcytosis, low serum iron, or low blood hemoglobin. Upon stratification by BOLA2 copy number, our data showed an association between low BOLA2 dosage and the above phenotypes (8/15 individuals with three copies, 53%, p ¼ 1e-4). In parallel, we analyzed hematological traits in mice carrying the 16p11.2 orthologous deletion or duplication, as well as Bola2 þ/À and Bola2 À/À animals. The Bola2-deficient mice and the mice carrying the deletion showed early evidence of iron deficiency, including a mild decrease in hemoglobin, lower plasma iron, microcytosis, and an increased red blood cell zinc-protoporphyrinto-heme ratio. Our results indicate that BOLA2 participates in iron homeostasis in vivo, and its expansion has a potential adaptive role in protecting against iron deficiency.

Maf1 −/− mice are lean, obesity-resistant and metabolically inefficient. Their increased energy expenditure is thought to be driven by a futile RNA cycle that reprograms metabolism to meet an increased demand for nucleotides stemming from the deregulation of RNA polymerase (pol) III transcription. Metabolic changes consistent with this model have been reported in both fasted and refed mice, however the impact of the fasting-refeeding-cycle on pol III function has not been examined. Here we show that changes in pol III occupancy in the liver of fasted versus refed wildtype mice are largely confined to low and intermediate occupancy genes; high occupancy genes are unchanged. However, in Maf1 −/− mice, pol III occupancy of the vast majority of active loci in liver and the levels of specific precursor tRNAs in this tissue and other organs are higher than wild-type in both fasted and refed conditions. Thus, MAF1 functions as a chronic repressor of active pol III loci and can modulate transcription under different conditions. Our findings support the futile RNA cycle hypothesis, elaborate the mechanism of pol III repression by MAF1 and demonstrate a modest effect of MAF1 on global translation via reduced mRNA levels and translation efficiencies for several ribosomal proteins. The synthesis of the translational apparatus is energetically costly and, consequently, subject to tight regulation (see 1 and references therein). For example, the activities of both pol I and III are controlled in response to nutrient availability 2-6. In the case of pol III, this control is exerted largely by the MAF1 protein, which constitutes its main repressor 7. In yeast, under favorable growth conditions, phosphorylation of Maf1 in a TOR complex 1-(TORC1) and protein kinase A-dependent manner inhibits its interaction with pol III and results in its nuclear exclusion 4,5. Upon nutrient deprivation or stresses like DNA damage, TORC1 is inactivated, Maf1 becomes dephosphorylated and shuttles into the nucleus where it represses pol III transcription. In mammalian cells, mTORC1 directly phosphorylates MAF1 on three residues (S60, S68, and S75) 8. Following serum deprivation, MAF1 undergoes dephosphorylation, increasingly targets pol III genes as determined by DamIP-seq, and represses pol III transcription 9,10. Mice lacking Maf1 are viable and fertile. They are slightly smaller and leaner than their wild-type (WT) counterparts, and they are resistant to diet-induced obesity and non-alcoholic fatty liver disease 11. This is due in part to reduced food intake but also to reduced metabolic efficiency. Maf1 −/− mice display increased energy expenditure throughout the diurnal cycle even though they are not physically more active than WT mice 11. Studies with fasted mice revealed elevated pol III transcription and precursor tRNA levels in the liver and numerous others tissues (~ 3 to 9 fold changes in precursor tRNAs) but no change in total tRNA or mature tRNA levels. These and other findings led to the proposal of a futile RNA cycle to account for t...

Maf1 -/mice are lean, obesity-resistant and metabolically inefficient. Their increased energy expenditure is thought to be driven by a futile RNA cycle that reprograms metabolism to meet an increased demand for nucleotides stemming from the deregulation of RNA polymerase (pol) III transcription. Metabolic changes consistent with this model have been reported in both fed and fasted mice, however the impact of the feeding-fasting cycle on pol III function has not been examined. Here we show that changes in pol III occupancy in the liver of fed versus fasted wild-type mice are largely confined to low and intermediate occupancy genes; high occupancy genes are unchanged. However, in Maf1 -/mice, pol III occupancy of the vast majority of active loci in liver and the levels of specific precursor tRNAs in this tissue and other organs are higher than wild-type in both fed and fasted conditions. Thus, MAF1 functions as a chronic repressor of active pol III loci and can modulate transcription under different conditions. Our findings support the futile RNA cycle hypothesis, elaborate the mechanism of pol III repression by MAF1 and demonstrate a modest effect of MAF1 on global translation via reduced mRNA levels and translation efficiencies for several ribosomal proteins. Affiliations

Human-specific duplications at chromosome 16p11.2 mediate recurrent pathogenic 600 kbp BP4-BP5 copy number variations, one of the most common genetic causes of autism. These copy number polymorphic duplications are under positive selection and include 3–8 copies of BOLA2, a gene involved in the maturation of cytosolic iron-sulfur proteins. To investigate the potential advantage provided by the rapid expansion of BOLA2, we assessed hematological traits and anemia prevalence in 379,385 controls and individuals who have lost or gained copies of BOLA2: 89 chromosome 16p11.2 BP4-BP5 deletion and 56 reciprocal duplication carriers in the UK Biobank. We found that the 16p11.2 deletion is associated with anemia (18/89 carriers, 20%, P=4e-7, OR=5), particularly iron-deficiency anemia. We observed similar enrichments in two clinical 16p11.2 deletion cohorts, with 6/63 (10%) and 7/20 (35%) unrelated individuals with anemia, microcytosis, low serum iron, or low blood hemoglobin. Upon stratification by BOLA2 copy number, we found an association between low BOLA2 dosage and the above phenotypes (8/15 individuals with three copies, 53%, P=1e-4). In parallel, we analyzed hematological traits in mice carrying the 16p11.2 orthologous deletion or duplication, as well as Bola2+/- and Bola2-/- animals. The deletion and Bola2-deficient mice showed early evidence of iron deficiency, including a mild decrease in hemoglobin, lower plasma iron, microcytosis, and an increased red blood cell zinc protoporphyrin to heme ratio. Our results indicate that BOLA2 participates in iron homeostasis in vivo and its expansion has a potential adaptive role in protecting against iron deficiency.