Abstract

Background

Toll-like receptors (TLRs) play a central role in innate immunity. TLRs are membrane glycoproteins and contain leucine rich repeat (LRR) motif in the ectodomain. TLRs recognize and respond to molecules such as lipopolysaccharide, peptidoglycan, flagellin, and RNA from bacteria or viruses. The LRR domains in TLRs have been inferred to be responsible for molecular recognition. All LRRs include the highly conserved segment, LxxLxLxxNxL, in which "L" is Leu, Ile, Val, or Phe and "N" is Asn, Thr, Ser, or Cys and "x" is any amino acid. There are seven classes of LRRs including "typical" ("T") and "bacterial" ("S"). All known domain structures adopt an arc or horseshoe shape. Vertebrate TLRs form six major families. The repeat numbers of LRRs and their "phasing" in TLRs differ with isoforms and species; they are aligned differently in various databases. We identified and aligned LRRs in TLRs by a new method described here.

Results

The new method utilizes known LRR structures to recognize and align new LRR motifs in TLRs and incorporates multiple sequence alignments and secondary structure predictions. TLRs from thirty-four vertebrate were analyzed. The repeat numbers of the LRRs ranges from 16 to 28. The LRRs found in TLRs frequently consists of LxxLxLxxNxLxxLxxxxF/LxxLxx ("T") and sometimes short motifs including LxxLxLxxNxLxxLPx(x)LPxx ("S"). The TLR7 family (TLR7, TLR8, and TLR9) contain 27 LRRs. The LRRs at the N-terminal part have a super-motif of STT with about 80 residues. The super-repeat is represented by STTSTTSTT or _TTSTTSTT. The LRRs in TLRs form one or two horseshoe domains and are mostly flanked by two cysteine clusters including two or four cysteine residue.

Conclusion

Each of the six major TLR families is characterized by their constituent LRR motifs, their repeat numbers, and their patterns of cysteine clusters. The central parts of the TLR1 and TLR7 families and of TLR4 have more irregular or longer LRR motifs. These central parts are inferred to play a key role in the structure and/or function of their TLRs. Furthermore, the super-repeat in the TLR7 family suggests strongly that "bacterial" and "typical" LRRs evolved from a common precursor.

No abstract

AimWe previously found that chronic tuberous sclerosis protein 2 (TSC2) deletion induces activation of mammalian target of rapamycin Complex 1 (mTORC1) and leads to hypertrophy of pancreatic beta cells from pancreatic beta cell-specific TSC2 knockout (βTSC2−/−) mice. The present study examines the effects of TSC2 ablation on insulin secretion from pancreatic beta cells.MethodsIsolated islets from βTSC2−/− mice and TSC2 knockdown insulin 1 (INS-1) insulinoma cells treated with small interfering ribonucleic acid were used to investigate insulin secretion, ATP content and the expression of mitochondrial genes.ResultsActivation of mTORC1 increased mitochondrial DNA expression, mitochondrial density and ATP production in pancreatic beta cells of βTSC2−/− mice. In TSC2 knockdown INS-1 cells, mitochondrial DNA expression, mitochondrial density and ATP production were increased compared with those in control INS-1 cells, consistent with the phenotype of βTSC2−/− mice. TSC2 knockdown INS-1 cells also exhibited augmented insulin secretory response to glucose. Rapamycin inhibited mitochondrial DNA expression and ATP production as well as insulin secretion in response to glucose. Thus, βTSC2−/− mice exhibit hyperinsulinemia due to an increase in the number of mitochondria as well as enlargement of individual beta cells via activation of mTORC1.ConclusionActivation of mTORC1 by TSC2 ablation increases mitochondrial biogenesis and enhances insulin secretion from pancreatic beta cells.