Glucocorticoid-Induced Cleft Palate in the Mouse: Two Major Histocompatibility Complex, H-2, Loci with Different Mechanisms

Isolated cleft palate is induced in the progeny of pregnant mice that are given glucocorticoids. The incidence varies among inbred strains and with dose and stage of gestation when the drug is given. One chromosomal region responsible for strain-associated differences in sensitivity is the major histocompatibility complex, H-2. H-2^a is associated with susceptibility, H-2^b with resistance. There appear to be both maternal and embryonic genetic factors affecting the sensitivity to glucocorticoids. In experiments reported here congenic strains of mice with H-2^a, H-2^d and H-2^k haplotypes on a C57BL/10 genomic background were used. This allowed the determination of the effect on sensitivity by two H-2 subregions; the subregions are H-2K to I-E and I-C to H-2D. Methods included dose-response analysis and reciprocal cross analysis using dexamethasone given on day 12 of pregnancy. Results show that each subregion affects the strain's sensitivity to dexamethasone-induced cleft palate. The regression coefficients for B10.A-H-2^a (45.4 ± 4.13) were different from those for B10.BR-H-2^k (67.2 ± 10.8) and B10.D2-H-2^d (70.5 ± 9.74). The estimated mean arcsine% cleft palate at 160 mg/kg was different for each strain: B10.A- H-2^a, 53.1 ± 2.19; B10.BR-H-2^k, 33.1 ± 2.27; B10.D2-H-2^d, 25.0 ± 2.75. Different patterns of change in sensitivity were observed among the reciprocal crosses. In summary, the H-2K to I-E subregion seemed to influence both maternal and embryonic factors, whereas only embryonic factors were influenced by the I-C to H-2D subregion. These data suggest that the mechanisms affecting glucocorticoid sensitivity which are genetically encoded within each H-2 subregion are different, and there is an interaction between the alleles. The mode of interaction can be either complementation or epistasis.     

Genetic control of human NK cell repertoire

Through differential killer cell Ig-like receptor (KIR) and CD94:NKG2 gene expression, human NK cells generate diverse repertoires, each cell having an inhibitory receptor for autologous HLA class I. Using a new method for measuring repertoire difference that integrates multiple flow cytometry parameters, we found individual repertoire stability, but population variability. Correlating repertoire differences with KIR and HLA genotype for 85 sibling pairs reveals the dominant influence of KIR genotype; HLA genotype having a subtle, modulating effect on relative KIR expression frequencies. HLA and/or KIR genotype also influences CD94:NKG2A expression. After HLA-matched stem cell transplantation, KIR repertoires either recapitulated that of the donor or were generally depressed for KIR expression. Human NK cell repertoires are defined by combinations of variable KIR and HLA class I genes and conserved CD94:NKG2 genes.     

TTK/hMps1 participates in the regulation of DNA damage checkpoint response by phosphorylating CHK2 on threonine 68

CHK2/hCds1 plays important roles in the DNA damage-induced cell cycle checkpoint by phosphorylating several important targets, such as Cdc25 and p53. To obtain a better understanding of the CHK2 signaling pathway, we have carried out a yeast two-hybrid screen to search for potential CHK2-interacting proteins. Here, we report the identification of the mitotic checkpoint kinase, TTK/hMps1, as a novel CHK2-interacting protein. TTK/hMps1 directly phosphorylates CHK2 on Thr-68 in vitro. Expression of a TTK kinase-dead mutant, TTK(D647A), interferes with the G(2)/M arrest induced by either ionizing radiation or UV light. Interestingly, induction of CHK2 Thr-68 phosphorylation and of several downstream events, such as cyclin B1 accumulation and Cdc2 Tyr-15 phosphorylation, is also affected. Furthermore, ablation of TTK expression using small interfering RNA results not only in reduced CHK2 Thr-68 phosphorylation, but also in impaired growth arrest. Our results are consistent with a model in which TTK functions upstream from CHK2 in response to DNA damage and suggest possible cross-talk between the spindle assembly checkpoint and the DNA damage checkpoint.     

Inhibition of voltage-gated K+ channels in dendritic cells by rapamycin

Rapamycin, an inhibitor of the serine/threonine kinase mammalian target of rapamycin (mTOR), is a widely used immunosuppressive drug. Rapamycin affects the function of dendritic cells (DCs), antigen-presenting cells participating in the initiation of primary immune responses and the establishment of immunological memory. Voltage-gated K(+) (Kv) channels are expressed in and impact on the function of DCs. The present study explored whether rapamycin influences Kv channels in DCs. To this end, DCs were isolated from murine bone marrow and ion channel activity was determined by whole cell patch clamp. To more directly analyze an effect of mTOR on Kv channel activity, Kv1.3 and Kv1.5 were expressed in Xenopus oocytes with or without the additional expression of mTOR and voltage-gated currents were determined by dual-electrode voltage clamp. As a result, preincubation with rapamycin (0-50 nM) led to a gradual decline of Kv currents in DCs, reaching statistical significance within 6 h and 50 nM of rapamycin. Rapamycin accelerated Kv channel inactivation. Coexpression of mTOR upregulated Kv1.3 and Kv1.5 currents in Xenopus oocytes. Furthermore, mTOR accelerated Kv1.3 channel activation and slowed down Kv1.3 channel inactivation. In conclusion, mTOR stimulates Kv channels, an effect contributing to the immunomodulating properties of rapamycin in DCs.     

Cw * 1701 defines a divergent African HLA-C allelic lineage

 The complete sequence of a new HLA-C allele, Cw ^* 1701, was determined from a South African Zulu individual. Unique features that distinguish Cw ^* 1701 from other HLA-C alleles include multiple point substitutions and an 18 nucleotide insertion in exon 5, which encodes the transmembrane domain. In a phylogenetic analysis of HLA-C sequences, Cw ^* 1701 forms a third, distinct allelic lineage. A comparison of the transmembrane domain of Cw ^* 1701 with other HLA-B and -C alleles reveals that duplications and deletions have been common in the evolution of these loci. A polymerase chain reaction based typing method was used to determine the distribution of this unusual allele in human populations. In contrast to the other two lineages of HLA-C alleles, the Cw ^* 17 lineage is found at high frequencies only in populations of African descent. In addition, the HLA-B/Cw ^* 17 haplotype diversity is higher in Africa. Received: 29 June 1996  / Revised: 20 December 1996     

Glucocorticoid-induced cleft palate genes in chromosome 17: Genetic linkage and mapping analyses

Genes that influence susceptibility to dexamethasone-induced cleft palate and tentatively designated Dcp are linked to the major histocompatibility complex H-2 in chromosome 17 of the mouse. Experiments presented refine the map of genes. The results show two or three Dcp loci. The two-locus model maps Dcp genes to the class II gene E and to the chromosomal region between the S and D genes. The three-locus model maps the Dcp genes to the chromosomal regions from the centromere to E , from E to S, and from D to Pgk-2. Experiments were done by comparing the dexamethasone-induced cleft palate dose response of congenic strains with H-2 haplotypes that are recombinants of H-2 ^aand H-2 ^b. The analysis of genetic linkage between H-2 and Dcp was expanded to include reciprocal backcrosses. A maternal factor was found to influence the frequency of dexamethasone-induced cleft palate in the backcross fetuses. The factor's origin is associated with the H-2 haplotype of the outcross mother, so the effect is actually a grandmother effect that probably is transmitted horizontally. Finally, the sexes were distributed unevenly between the fetuses with cleft palate in two of the congenic strains. This suggests interaction between the H-2-linked Dcp genes and a Dcp sex-associated gene that modulates susceptibility to dexamethasone-induced cleft palate.