It is not clear how DNA hypomethylation in ICF patients may result in the immunodeficiency. However, it seems likely, as described under "Etiology," that it is the DNA methylation deficiency, and not some other aspect of impaired DNMT3B activity, that is responsible for the disease. Hypomethylation in some region(s) of the genome probably changes transcription of one or more genes that are primary targets for the deficiency in DNMT3B activity.
The immunodeficiency in ICF is manifested as low serum immunoglobulin levels, although there can be normal B- and T-cell numbers in peripheral blood. The latter finding indicates that the early stages of lymphocyte differentiation are not abnormal. High percentages of cells in ICF B-cell lines that have membrane-bound IgM and/or IgD have been observed even in patients with extremely low levels of IgM. Therefore, class switching was normal for the precursors of these cells, but there seems to be a defect in lymphocyte maturation or activation [
14]. In a comparison of ICF lymphoblastoid cell lines (B-cell lines with known
DNMT3B mutations) and analogous control cell lines by a microarray expression analysis, only a small number of genes, including IgG- and IgA-encoding sequences, were found to have ICF-specific differences in RNA levels [
14]. A larger microarray expression analysis, which was followed by quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR) for selected genes, gave similar findings (M. Ehrlich, C. Sanchez, C. Shao, R. Kuick, S. Hanash, unpub. data). In the first study, no ICF-specific differences in promoter methylation were seen, even for genes with elevated RNA levels in ICF cell lines [
14] and similar methylation analyses are underway for selected genes from the second study. A number of reports of promoter-region hypomethylation in ICF cells, including in the inactive X chromosome, have revealed inconsistent hypomethylation at given gene regions among different patients [
32,
36,
64,
65]. This finding matches results from a genome-wide search for consistently hypomethylated DNA sequences in ICF
vs. control B-cells lines by 2-dimensional gel electrophoresis of DNA digested with two restriction endonucleases, one of which was CpG methylation-sensitive [
66]. Only repeated DNA sequences (a subtelomeric repeat and a repeat from the acrocentric chromosomes) were identified as hypomethylated specifically in all the ICF samples by this analysis. It has been hypothesized that in ICF lymphoid cells, hypomethylation of regions of the genome that are normally constitutively heterochromatic (
e.g., 1qh and 16qh) could affect regulation of expression of genes elsewhere in the genome by altering the postulated normal sequestration of DNA sequence-specific proteins at this heterochromatin [
14,
26]. There are precedents for such binding of transcription factors to centromeric heterochromatin [
67,
68]. The hypothesis that constitutive heterochromatin itself can act in
trans to modulate gene expression remains to be tested for ICF cells. Alternatively, there may be only a small number of currently unidentified gene regions with consistent hypomethylation specific to ICF lymphoid cells that are responsible for ICF-type immune dysfunction.