By David N. Leff

Overly devout sun worshipers suffer the slings and arrows of aggressive ultraviolet (UV) radiation, which inflicts insults ranging from sunburn to skin cancer. People overexposed to ionizing radiation - mainly diagnostic and therapeutic X-rays (not to mention nuclear weapons testing) - risk various cancers, notably tumors of the immune system's B and T cells, which cause leukemias and lymphomas.

These neoplasms usually arise from fractures across the double-stranded helix of DNA sequences in vital genes. For example, a once-rare, life-threatening skin cancer called xeroderma pigmentosum - now common in AIDS victims - results from the lack of a protective enzyme called UV-specific endonuclease. This is only one of a small army of caretaker proteins in the body, trained to repair damaged DNA.

Interestingly, double-stranded genomic breaks also occur by nature's design as well as by carcinogenic mishap. These benign DNA cross-cuts happen in the course of the immune system's endless rearrangement of genes encoding proteins that perform the intricate ballet dance by which its B cells generate the more than 100 million disparate antibodies poised to attack any and all possible infectious antigens the flesh may be heir to. This process, called V(D)J recombination, involves the genes that encode the variable (V), diversity (D) and joining (J) amino-acid sequences that make up the future antibody's antigen-spotting nose cone.

To knit up the raveled sleeves of DNA double breaks, a squadron of special caretaker molecules musters non-homologous end-joining proteins (NHEJ), of which the most recently identified is XRCC4. This protein was discovered by immunologist Frederick Alt, of Harvard Medical School in Boston - a pioneer in researching the hyper-devious processes of antibody formation and immune system cancers.

Alt, also a Howard Hughes Medical Institute investigator, is senior author of a paper in the current issue of Nature, dated April 20, 2000. Its title: "Interplay of p53 and DNA-repair protein XRCC4 in tumorigenesis, genomic stability and development."

"Our findings," the article sums up, "support a crucial role for the non-homologous end-joining pathway as a caretaker of the mammalian genome, a role required for both normal development and for suppression of tumors."

Alt told BioWorld Today, "The way we started studying this particular set of proteins and the non-homologous end-joining reaction, came from our prior studies of V(D)J recombination, by which lymphocytes assemble gene segments and encode their antigen receptor, to make antibody genes for example."

Cutting, Pasting, Broken DNA Strands

"What happens in lymphocytes when they're developing," Alt continued, "is that a very specific set of enzymes cut these gene segments, so they can then be pasted back together. This XRCC4 belongs to a set of proteins that are involved not in specific strand-cutting, but rather in the pasting back together. So in the absence of those caretaker proteins, the developing lymphocytes can't make their antigen receptor, or immunoglobulin [antibody] gene, simply because they can't reunite those gene segments."

Alt described a series of in vivo experiments focusing on a mouse that lacked both XRCC4 genes. "It regenerated," he recounted, "but with a resulting major defect, the fact that its lymphocyte development was completely impaired. If the cells make the DNA breaks in the absence of XRCC4, they can't put the ends back together. So, No. 1, they have broken DNA, from which they would die by normal surveillance mechanisms; and No. 2, they can't make the genes, so they don't make immunoglobulin proteins that are necessary to promote their development and survival. Thus, their immune systems were blocked.

"When we eliminated both XRCC4 genes," he went on, "we found additional effects on the mice. One was that they died late in embryonic gestation, and when we looked we found that throughout the brains of these animals, the recently developed neurons were dying from apoptosis - programmed cell suicide. To try to understand this process, we reasoned that the apoptotic death is likely being affected or signaled by one of the common DNA surveillance proteins. The likely candidate was the p53 tumor suppressor, and genome guardian, which can recognize DNA breaks and signal cells to arrest their cell-division cycle, so they could have time to repair them. If this failed and nothing else could be done, p53 would signal the cells to die, so that they wouldn't go on living with all sorts of broken DNA."

What Alt and his co-authors then did was introduce p53 deficiency into their XRCC4-deficient mice, which completely rescued them from the lethality of the XRCC4-minus mutations. "Meaning," Alt recounted, "that the mice no longer died embryonically, but were born, lived, and seemed fairly normal. And when we looked at their nervous systems we no longer saw this substantial cell death, so our notion that it could be a p53-dependent process going to neurons proved to be correct. However, their immune systems didn't develop, and they were still blocked for that."

Going on with the in vivo account, Alt recalled that "the mice now survived, but while they were still quite young, they all developed a very aggressive progenitor-B-cell lymphoma. And that was notable because p53-deficient mice also developed cancer, but at a much later age, and with a different type of tumor - T-cell lymphoma."

On Trail Of Anonymous Caretaker

"The DNA end-joining process," he observed, "recognizes those broken ends and normally puts them back together. But if you don't have that particular caretaker protein, then presumably another - as yet unidentified - repair pathway will fix them. So what we would see by looking at these lymphoid tumors, in these mice, this other pathway can frequently lead to adverse chromosomal translocations."

Alt and his team now plan "first of all, in the context of the lymphomas, to look in detail at the DNA break points, to understand what - in the absence of end-joining - that alternative mechanism is that causes chromosomal translocation of the immunoglobulin heavy chain locus to the c-myc [tumorigenic oncogene] locus.