An inherited failure of gene regulation

The class II molecules of the major histocompatibilty complex (MHC) are involved in presenting antigens to CD4 T cells. The peptide antigens that they present are derived from extracellular pathogens and proteins taken up into intracellularly vesicles, or from pathogens such as Mycobacterium that persist intracellular inside vesicles. MHC class II molecules are expressed constitutively on antigen-presenting cells, including B lymphocytes, macrophages, and dendritic cells. In humans, together with the MHC class I molecules they are known as the HLA antigens. They are also expressed on the epithelial cells of the thymus and their expression can be induced on other cells, principally by the cytokine interferon -?. T cells also express MHC class II molecules when they are activated.

MHC class II molecules are heterodimers consisting of an a chain and a ß chain. The genes encoding both chains are located in the MHC on the short arm of chromosome 6 in humans. The principal MHC class II molecules are designated DP, DQ and DR, and like the MHC class I molecules they are highly polymorphic. Peptides bound to MHC class II molecules can be recognized only by the T-cell receptors of CD4 T cells and not by those of CD8 T cells. MHC class II molecules expressed in the thymus also have a vital role in the intrathymic maturation of CD4 T cells.

Expression of the genes encoding the a and ß chains of MHC class II molecules must be coordinated strictly and is under complex regulatory control. The regulation of MHC class II gene expression is not fully understood as it involves the action of transcription factors that are defined only in part. The existence of these transcription factors and a means of identifying then were first suggested by the study of patients with MHC class II deficiency.

The case of Helen Burns: a 6-month-old child with a mild form of severe combined immunodeficiency.

Helen Burns was the second child born to her parents. She thrived until 6 months of age when she developed pneumonia in both lungs, accompanied by a severe cough and fever. Blood and sputum cultures for bacteria were negative but a tracheal aspirate revealed the presence of abundant Pneumocystis carinii . She was treated successfully with the anti- Pneumocystis drug pentamidine and seemed to recover fully.

As her pneumonia was caused by the opportunistic pathogen Pneumocystis carinii , Helen was suspected to have severe combined immunodeficiency. A blood sample was taken and her peripheral blood mononuclear cells were stimulated with phytohemagglutinin (PHA) to test for T cells function by 3 H-thymidine incorporation into DNA. A normal T-cell proliferative response was obtained, with her T cells incorporating 114,020 counts min -1 of 3 H-thymidine (normal control 75,000 counts min- 1 ). Helen had received routine immunizations with orally administrated polio vaccine and DPT (diphtheria, pertussis, and tetanus) vaccine at 2 months old. However, in further tests, her T cells failed to respond to tetanus toxin in vitro , although they responded normally in the 3 H-thymidine incorporation assay when stimulated with allogeneic B cells (6730 counts min- 1 incorporated compared with 783 counts min- 1 for unstimulated cells.

When it was found that Helen's T cells could not respond to a specific antigenic stimulus, her serum immunoglobulins were measured and found to be very low. IgG levels were 96 mg dl- 1 (normal 600-1400 mg dl- 1 ), IgA was 6 mg dl- 1 (normal 60-380 mg dl- 1 ), and IgM 30 mg dl- 1 (normal 40-345 mg dl- 1 ).

Helen's white blood cell count was elevated at 20,000 cells µl- 1 ). (normal range 4000-7000 µl- 1 ) Of these, 82% were neutrophils, 10% lymphocytes, 6% monocytes, and 2% eosinophils. The calculated number of 2000 lymphocytes µl- 1 is low for her age (normal >3000 µl- 1 ). Of her lymphocytes, 7% were B cells as determined by an antibody to CD20 (normal 10-12%) and 57% reacted with antibody to the T cell marker CD3. Of these T cells, 34% were positive for CD8, and 20% were positive for CD4). Thus, at 388 cells µl- 1 her number of CD8 T cells was within the normal range, but the number of CD4 T cells (288 µl- 1 ) was much lower than the normal (her CD4 T-cell count would be expected to be twice her CD8 T-cell count). The presence of substantial numbers of T cells, and thus a normal response to PHA, ruled out a diagnosis of sever combined immunodeficiency.

Helen's pediatrician referred her to the Children's Hospital for consideration for a bone marrow transplant, despite the lack of diagnosis. When an attempt was made to HLA type Helen, her parents and her healthy 4-year-old brother, a DR type count not be obtained from Helen's white blood cells. A long-term culture of her B cells was made by transforming them with Epstein-Barr virus and the transformed B cells were then examined for expression of MHC class I and class II molecules with fluorescent-tagged antibodies. It was found that her B cells did not express HLA-DQ or HLA-DR molecules and a diagnosis of MHC class II deficiency was established.

As her brother did not have the same HLA type as Helen, it was decided to use her mother as a bone marrow donor. Helen was given mg kg- 1 body weight of cytotoxic drug busulfan every 6 hours for 4 days to depress bone marrow function and then 50 mg kg- 1 of cyclophosphamide to ablate her bone marrow. The maternal bone marrow was depleted of T cells to diminish the chance of graft-versus-host disease developing and was administered to Helen by transfusion. The graft was successful and immune function was restored.

MHC class II deficiency

MHC class II deficiency is inherited as an autosomal recessive trait. Health problems showed up early in infancy. Affected babies present the physician with a mild form of severe combined immunodeficiency (SCID) as they have increased susceptibility to pyogenic and opportunistic infections. However, they differ from SCID patients in that they have T cells, which can respond to nonspecific T-cell mitogens such as PHA and to allogeneic stimuli. Also, unlike SCID patients, they do not sustain graft-versus-host disease when given HLA-mismatched blood transfusions. This is because the host tissue has no MHC class II molecules against which the T cells in the graft can react. SCID patients, in contrast, carry MHC II molecules on some of their tissues, against which T cells in the graft will react to cause graft-versus-host disease. They have no T cells at all and therefore cannot reject the graft. Unlike in some other types of immunodeficiency, progressive infection with the attenuated live vaccine strain BCG has not been observed in MHC class II-deficient patients after BCG vaccination against tuberculosis (most cases of MHC class II deficiency have been observed in North African migrants in Europe, where BCG vaccination is routine). This is because mycobacterial antigens derived from BCG can be presented on MHC class I molecules and infected cells can be destroyed by cyotoxic T cells.

Patients with MHC class II deficiency are deficient in CD4 T cells, in contrast to MHC class I deficiency, in which CD8 T cells numbers are very low and the levels of CD4 cells are normal. They also have moderate to severe hypogammaglobulinemia.

Genetic linkage analysis in large extended families with MHC class II deficiency has shown that this condition is not linked to the MHC locus on the short arm of chromosome 6 and that the genes encoding the MHC class II molecules at this locus are normal. Interferon -g induces the expression of MHC class II molecules on antigen-presenting cells from normal people but fails to induce their expression on the antigen-presenting cells of patients with MHC class II deficiency. This suggested that the defect might lie in the regulation of expression of the MHC class II genes.

The search for the cause of the defect was complicated further by the discovery that MHC class II deficiency in different patients seems to have different causes. B-cell lines isolated from class II-deficient patients do not express MHC class II molecules. However, when B cells from two different patients are fused, MHC class II expression is often observed. The fusion of the two cell lines has corrected the defect. This means that one cell must be able to replace whatever is lacking in the other, and thus the two cells must carry different genetic defects causing the MHC class II deficiency. Pairwise fusions were performed on a large number of cell lines from different patients, and at least four complementation groups were found.

These experiments provided clues that led eventually to the identification of the defect. The lack of MHC class II molecules turns out to result from defects in the transcriptions factors required to regulate their coordinated expression. All four of these transcriptions factors, which bind to the 5' regulatory region of the MHC class II genes, have been identified.

We would like to thank Dr. Fred Rosen and Dr. Raif Geha for their contribution of the above information from their book, "Case Studies in Immunology 3."

Back to