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Tristetraprolin/ZFP36 Regulates the Turnover of Autoimmune-Associated HLA-DQ mRNAs

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Tristetraprolin/ZFP36 Regulates the Turnover of Autoimmune-Associated HLA-DQ mRNAs

Laura Pisapia et al. Cells.

Abstract

HLA class II genes encode highly polymorphic heterodimeric proteins functioning to present antigens to T cells and stimulate a specific immune response. Many HLA genes are strongly associated with autoimmune diseases as they stimulate self-antigen specific CD4+ T cells driving pathogenic responses against host tissues or organs. High expression of HLA class II risk genes is associated with autoimmune diseases, influencing the strength of the CD4+ T-mediated autoimmune response. The expression of HLA class II genes is regulated at both transcriptional and post-transcriptional levels. Protein components of the RNP complex binding the 3'UTR and affecting mRNA processing have previously been identified. Following on from this, the regulation of HLA-DQ2.5 risk genes, the main susceptibility genetic factor for celiac disease (CD), was investigated. The DQ2.5 molecule, encoded by HLA-DQA1*05 and HLA-DQB1*02 alleles, presents the antigenic gluten peptides to CD4+ T lymphocytes, activating the autoimmune response. The zinc-finger protein Tristetraprolin (TTP) or ZFP36 was identified to be a component of the RNP complex and has been described as a factor modulating mRNA stability. The 3'UTR of CD-associated HLA-DQA1*05 and HLA-DQB1*02 mRNAs do not contain canonical TTP binding consensus sequences, therefore an in silico approach focusing on mRNA secondary structure accessibility and stability was undertaken. Key structural differences specific to the CD-associated mRNAs were uncovered, allowing them to strongly interact with TTP through their 3'UTR, conferring a rapid turnover, in contrast to lower affinity binding to HLA non-CD associated mRNA.

Keywords: Human Leukocyte Antigen (HLA), RNA binding protein; RNA stability; RNA structure; celiac disease.

Conflict of interest statement

The authors declare no conflict of interest

Figures

Figure 1
Figure 1
RNA binding proteins interaction. (A) REMSA experiments performed using 3DQA101 (Lane 1) and 3DQA105 (Lane 9) riboprobes. Lanes 2 and 8 show the digestion of riboprobes with T1 RNase. The binding of DQA101 with M14 extract is in Lane 3 and with B-LCL#5 extract in Lane 6. The binding of 3DQA105 with M14 extract is in Lane 9 and with B-LCL#5 extract in Lane 12. The interaction of 5 and 1 µg of rTTP with 3DQA101 is shown in Lanes 4 and 5 and with 3DQA105 in Lanes 10 and 11. (B) Western blot analysis of biotin pull-down assay carried by using 3DQA101, 3DQA105, 3DQB102, and 3DQB105 riboprobes. The antibodies used for the immunoblot were anti-NF90, anti-TTP, and anti-EBP1. Molecular weights are as indicated.
Figure 2
Figure 2
Sequences and structures comparison between 3′UTR of DQA1* and DQB1* genes. (A) Comparison of the 3′UTR sequences for the DQA1*01 and DQA1*05 alleles indicates 94.1% sequence identity. There is a slightly higher proportion of GC content in the DQA1*01 allele (0.46) as compared to DQA1*05 (0.44). (B) The DQB1*05 and DQB1*02 alleles show a lower sequence identity (92.3%), but identical GC proportions (0.55). (C,D) RNA secondary structure prediction reveals the sequence differences between alleles impacts the minimum-free energy folding (RNAfold). Common structural motifs from FOLDALIGN are mapped onto the structures as well as canonical ARE (AUUUA/AUUUUA) and half-ARE motifs (e.g., UAUU).
Figure 3
Figure 3
AU-rich motif analysis of the DQA1* and DBQ1* genes reveals key differences. (A) All possible di-, tri-, tetra-, penta- and hexa-mers are assessed for their observed frequencies against their expected frequencies. AU-rich motifs are colored, non-AU rich in grey. All AU-rich kmer motifs have higher enrichments in the CD associated DQA1*05 and DQB1*02 alleles as compared to the non-CD associated DQA1*01 and DQB1*05 alleles. (B) Mapping the AU-rich motifs to their position within the 3′UTR sequences reveals a clear enrichment in AU-motifs throughout the CD associated alleles DQA1*05 and DQB1*02 (Blue) as compared to the non-CD associated DQA1*01 and DQB1*05 alleles (Red). Nested motifs are indicated on the y-axis; for example, two overlapping motifs will show a score of 2. A structure accessibility score indicates the likelihood the RNA single stranded, an absence of base-pairing, across its length. A score of 1.0 indicates the structure is single stranded.
Figure 4
Figure 4
TTP knockdown in M14. (A) Western blot performed with anti-TTP for the assessment of protein depletion 48 h after silencing with siCtlr or siTTP (B) Fold variation of DRA, DRB1, and DQA1 mRNAs. HLA-a,b,c are the class I mRNAs, CIITA is the HLA class II transcriptional transactivator.
Figure 5
Figure 5
TTP knockdown in B-LCL#5. (A). Western blot performed with anti-TTP for the assessment of protein depletion 48 h after silencing with siCtlr or siTTP nucleofection. (B) Cytofluorimetric analysis of HLA-DQ surface expression, reported as fold change of MFI (Mean Fluorescence Intensity). (C) Fold variation of DRA, DRB1, DQA1, DQB1, and HLA-A, -B, and -C (class I) mRNAs. (D) Fold variation of DQA1*01, DQA1*05, DQB1*02, and DQB1*05 mRNAs.

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References

    1. Ting J.P.Y., Trowsdale J. Genetic control of MHC class II expression. Cell. 2002;109:S21–S33. doi: 10.1016/S0092-8674(02)00696-7. - DOI - PubMed
    1. Reith W., LeibundGut-Landmann S., Waldburger J.M. Regulation of MHC class II gene expression by the class II transactivator. Nat. Rev. Immunol. 2005;5:793–806. doi: 10.1038/nri1708. - DOI - PubMed
    1. Corso C., Pisapia L., Citro A., Cicatiello V., Barba P., Cigliano L., Abrescia P., Maffei A., Manco G., Del Pozzo G. EBP1 and DRBP76/NF90 binding proteins are included in the major histocompatibility complex class II RNA operon. Nucleic Acids Res. 2011;39:7263–7275. doi: 10.1093/nar/gkr278. - DOI - PMC - PubMed
    1. Pisapia L., Cicatiello V., Barba P., Malanga D., Maffei A., Hamilton R.S., Del Pozzo G. Co-regulated expression of alpha and beta mRNAs encoding HLA-DR surface heterodimers is mediated by the MHCII RNA operon. Nucleic Acids Res. 2013;41:3772–3786. doi: 10.1093/nar/gkt059. - DOI - PMC - PubMed
    1. Ko H.R., Chang Y.S., Park W.S., Ahn J.Y. Opposing roles of the two isoforms of ErbB3 binding protein 1 in human cancer cells. Int. J. Cancer. 2016;139:1202–1208. doi: 10.1002/ijc.30165. - DOI - PubMed

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