Epididymis-specific RNase A family genes regulate fertility and small RNA processing.

Sperm small RNAs are implicated in intergenerational transmission of paternal environmental effects. Small RNAs generated by cleavage of tRNAs, known as tRNA fragments (tRFs), are an abundant class of RNAs in mature sperm, and can be modulated by environmental conditions. The ribonuclease(s) responsible for the biogenesis of tRFs in the male reproductive tract remains unknown. Angiogenin, a member of the Ribonuclease A superfamily (RNase A), cleaves tRNAs to generate tRFs in response to cellular stress. Four paralogs of Angiogenin, namely Rnase9, Rnase10, Rnase11, and Rnase12, are specifically expressed in the epididymis-a long, convoluted tubule where sperm mature and acquire fertility and motility. The biological functions of these genes remain largely unknown. Here, by generating mice deleted for all four genes (Rnase9-12-/-, termed "KO" for Knock Out), we report that these genes regulate fertility and RNA processing. KO mice showed complete male sterility. KO sperm fertilized oocytes in vitro but failed to efficiently fertilize oocytes in vivo, likely due to an inability of sperm to pass through the utero-tubular junction. Intriguingly, there were decreased levels of fragments of tRNAs (tRFs) and rRNAs (rRNA-derived small RNAs or rsRNAs) in the KO epididymis and epididymal luminal fluid, implying that Rnase9-12 regulate the biogenesis and/or stability of tRFs and rsRNAs. Importantly, KO sperm showed a dramatic decrease in the levels of tRFs, demonstrating a role of Rnase9-12 in regulating sperm RNA composition. Together, our results reveal an unexpected role of four epididymis-specific non-canonical RNase A family genes in fertility and RNA processing.

Acetylcarnitine shuttling links mitochondrial metabolism to histone acetylation and lipogenesis.

The metabolite acetyl-CoA is necessary for both lipid synthesis in the cytosol and histone acetylation in the nucleus. The two canonical precursors to acetyl-CoA in the nuclear-cytoplasmic compartment are citrate and acetate, which are processed to acetyl-CoA by ATP-citrate lyase (ACLY) and acyl-CoA synthetase short-chain 2 (ACSS2), respectively. It is unclear whether other substantial routes to nuclear-cytosolic acetyl-CoA exist. To investigate this, we generated cancer cell lines lacking both ACLY and ACSS2 [double knockout (DKO) cells]. Using stable isotope tracing, we show that both glucose and fatty acids contribute to acetyl-CoA pools and histone acetylation in DKO cells and that acetylcarnitine shuttling can transfer two-carbon units from mitochondria to cytosol. Further, in the absence of ACLY, glucose can feed fatty acid synthesis in a carnitine responsive and carnitine acetyltransferase (CrAT)-dependent manner. The data define acetylcarnitine as an ACLY- and ACSS2-independent precursor to nuclear-cytosolic acetyl-CoA that can support acetylation, fatty acid synthesis, and cell growth.

Targeting ACSS2 with a Transition-State Mimetic Inhibits Triple-Negative Breast Cancer Growth.

Acetyl-CoA is a vitally important and versatile metabolite used for many cellular processes including fatty acid synthesis, ATP production, and protein acetylation. Recent studies have shown that cancer cells upregulate acetyl-CoA synthetase 2 (ACSS2), an enzyme that converts acetate to acetyl-CoA, in response to stresses such as low nutrient availability and hypoxia. Stressed cancer cells use ACSS2 as a means to exploit acetate as an alternative nutrient source. Genetic depletion of ACSS2 in tumors inhibits the growth of a wide variety of cancers. However, there are no studies on the use of an ACSS2 inhibitor to block tumor growth. In this study, we synthesized a small-molecule inhibitor that acts as a transition-state mimetic to block ACSS2 activity in vitro and in vivo. Pharmacologic inhibition of ACSS2 as a single agent impaired breast tumor growth. Collectively, our findings suggest that targeting ACSS2 may be an effective therapeutic approach for the treatment of patients with breast cancer. SIGNIFICANCE: These findings suggest that targeting acetate metabolism through ACSS2 inhibitors has the potential to safely and effectively treat a wide range of patients with cancer.