NoBlood

Blood, 15 November 2003, Vol. 102, No. 10, pp. 3556-3561.
Prepublished online as a Blood First Edition Paper on July 31, 2003; DOI 10.1182/blood-2003-05-1537.

HEMATOPOIESIS

Defective RNA ribose synthesis in fibroblasts from patients with thiamine-responsive megaloblastic anemia (TRMA)

László G. Boros, Mara P. Steinkamp, Judith C. Fleming, Wai-Nang Paul Lee, Marta Cascante and Ellis J. Neufeld

From the Harbor-University of California, Los Angeles (UCLA) Research and Education Institute, UCLA School of Medicine, Torrance, CA; the Division of Hematology/Oncology, Children's Hospital, Boston, MA; the Dana Farber Cancer Institute and Harvard Medical School, Boston, MA; and the Department of Biochemistry and Molecular Biology, University of Barcelona, Barcelona, Catalonia, Spain.

Fibroblasts from patients with thiamine-responsive megaloblastic anemia (TRMA) syndrome with diabetes and deafness undergo apoptotic cell death in the absence of supplemental thiamine in their cultures. The basis of megaloblastosis in these patients has not been determined. Here we use the stable [1,2-13C2]glucose isotope-based dynamic metabolic profiling technique to demonstrate that defective high-affinity thiamine transport primarily affects the synthesis of nucleic acid ribose via the nonoxidative branch of the pentose cycle. RNA ribose isolated from TRMA fibroblasts in thiamine-depleted cultures shows a time-dependent decrease in the fraction of ribose derived via transketolase, a thiamine-dependent enzyme in the pentose cycle. The fractional rate of de novo ribose synthesis from glucose is decreased several fold 2 to 4 days after removal of thiamine from the culture medium. No such metabolic changes are observed in wild-type fibroblasts or in TRMA mutant cells in thiamine-containing medium. Fluxes through glycolysis are similar in TRMA versus control fibroblasts in the pentose and TCA cycles. We conclude that reduced nucleic acid production through impaired transketolase catalysis is the underlying biochemical disturbance that likely induces cell cycle arrest or apoptosis in bone marrow cells and leads to the TRMA syndrome in patients with defective high-affinity thiamine transport. (Blood. 2003;102: 3556-3561)

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