TLR8 escapes X chromosome inactivation in human monocytes and CD4⁺ T cells.

"our r scripts performing two- and three-group meta-analytical summarization hypothesis tests and plotting are available from the zenodo repository under digital object identifiers (dois) https://doi org/10 5281/zenodo 6580369 and https://doi org/10 5281/zenodo 6580378 .; r scripts developed for the analysis of rna fish data are publicly available in the zenodo repository and the relevant identifiers are provided in the materials and methods section. to test for independence in 3 x 2 tables encompassing sparse cells (additional file 1 : s5 data) we performed ?; as described earlier for monocytes we concluded to xci escape in cd4 + t cells from women based on the presence of tlr7 (additional file 2 : fig s3d) or tlr8 (additional file 2 : fig s3e) transcripts on both x chromosomes of a cell or alternatively on identifying a single transcriptional signal on the xi (not shown) tlr7 evaded xci in 5% to 11% of t cells with a group mean of 8% (fig 2 g and additional file 1 : s1 data).; for tlr8 the frequency of escape cells varied over a 4% to 20% range with a group mean of 8% (fig 2 h and additional file 1 : s1 data).; as with female monocytes we were able to establish xci escape in all the individuals of this study group based on transcriptional tlr8 or tlr7 foci present on both x chromosomes (fig 3 a top and bottom right) or on a single signal ascribable to the xi (fig 3 a top and bottom left) tlr7 evaded xci in 10% to 17% of ks male monocytes (a narrower range than in women) with a group mean of 13% (fig 3 b and additional file 1 : s1 data).; the analysis for tlr7 and tlr8 transcripts in cd4 + t cells circumscribed to women and xy men showed a sex bias in the same direction as in monocytes (additional file 2 : fig s6a and s6b additional file 1 : s6 data).; descriptive and analytical statistics were computed from each table (additional file 1 : s3 data) and summarized group-wise by meta-analysis.; to investigate this possibility we cross-classified the cell counts in the rna fish data as2 x 2 contingency tables i e monocytes were stratified depending on the presence of signals for both tlr7 and tlr8 either gene alone or neither gene (additional file 1 : s3 data).; the preceding analyses scored the cells without regard to the chromosome of origin of the transcripts in women and ks men ("any x" data) but we carried out a parallel scoring procedure (additional file 1 : s4 data) restricted to xa + cells and considering only the alleles on the xa.; because only a fraction of the female monocytes analyzed exhibited xci escape the cross-classified cell counts based only on the xi signals encompassed fewer positive cells with instances of zero events in the double-positive (tlr7 + tlr8 + ) stratum (additional file 1 : s2 data).; to analyze these sparse data for yule's q we pooled the cell counts group-wise to increase statistical power (additional file 1 : s2 data).; next we contrasted these tlr7-tlr8 co-transcription data for the xi with the data for the xa of women and ks men (fig 5 e f and additional file 1 : s5 data).; the tests demonstrated significant differences between the xa and the xi for all individuals (additional file 1 : s5 data) and there is therefore a conclusive divergence between the marked tlr7-tlr8 transcriptional co-dependency on the xa (fig 5 e f ) and the trend for non-dependency observed on the xi alleles (additional file 2 : fig s7a b).; we scored first the cells regardless of the xa marking (any x data) and fig 6 a shows that similar to monocytes tlr7 + tlr8 + events among female t cells were more frequent than among the male cells: 13% of cells versus only 2% in males a 6:1 ratio consistent with the 7:1 ratio observed earlier in monocytes (additional file 1 : s6 data).; pcr primer pairs used in the preparation of the rna fish probes s1 data tlr7 escape from xci (monocytes women) s2 data.; donor ids in red s3 data.; donor ids in red s4 data.; donor ids in red s5 data.; donor ids in red s6 data."

"our r scripts performing two- and three-group meta-analytical summarization hypothesis tests and plotting are available from the zenodo repository under digital object identifiers (dois) https://doi org/10 5281/zenodo 6580369 and https://doi org/10 5281/zenodo 6580378 .; r scripts developed for the analysis of rna fish data are publicly available in the zenodo repository and the relevant identifiers are provided in the materials and methods section."

"Declarations Ethics approval and consent to participateThis study was approved by the relevant ethics board as described in the Materials and methods section. Written consent was obtained from each patient or, for child participants, from the legal guardian. Consent for publicationNot applicable. Competing interestsThe authors declare that they have no competing interests. Competing interests The authors declare that they have no competing interests."

"Funding This work was supported by grants from the Scleroderma Foundation to FJB and JCG, and from the FOREUM Foundation for Research in Rheumatology to JCG. JCG is supported by further grants from the Agence Nationale de la Recherche (ANR-20-CE15-0014; ANR-20-COV8-0004-01), Fondation pour la Recherche Médicale (DEQ20180339187), the Inspire Program from Région Occitanie/Pyrénées-Méditerranée (#1901175), and the European Régional Development Fund (MP0022856). BFL was supported by a post-doctoral fellowship from the Inspire Program from Région Occitanie/Pyrénées-Méditerranée (#1901175). AY was supported by a fellowship from SIDACTION, CSL Behring Research funds, an award from Fondation des Treilles, and a bursary from Association de la Charité des Jeunes de Kafarsir (Lebanon)."