Purine metabolism during neuronal differentiation: the relevance of purine synthesis and recycling

Abstract

Purines are a class of small organic molecules that are essential for all cells. They play critical roles in neuronal differentiation and function. Their importance is highlighted by several inherited disorders of purine metabolism, such as the Lesch-Nyhan disease, which is caused by a deficiency of the purine salvage enzyme, hypoxanthine-guanine phosphoribosyltransferase (HGprt). Despite the known importance of purines in the nervous system, knowledge regarding their metabolism in neurons is limited. In the current studies, purine pools and their metabolism were examined in rat PC6-3 cells, a PC12 pheochromocytoma subclone that undergoes robust differentiation with nerve growth factor. The results were compared with five new independent PC6-3 subclones with defective purine recycling due to different mutations affecting HGprt enzyme activity. The results demonstrate an increase in most purines and in energy state following neuronal differentiation, as well as specific abnormalities when purine recycling is lost. The loss of HGprt-mediated purine recycling also is associated with significant loss of dopamine and related metabolites in the mutant PC6-3 lines, suggesting an important connection between purine and dopamine pathways. These results provide insights into how purine pools and metabolism change with neuronal differentiation, and how specific enzyme defects may cause neuronal dysfunction.

Transcriptomic Approach to Lesch-Nyhan Disease

“…Interestingly, prior studies have shown that HPRT-deficient neuroblastoma subclones grow more slowly in culture than the parent cell from which they were derived. [22,23] Moreover, dysregulation of cell-cycle/cell-division genes also could play an important role in brain development: Cdk1, we found significantly downregulated in LND cells, has been linked to cell death of postmitotic neurons in brain development [24] and the Cdk1 complex plays a prime role in regulating N-myc phosphorylation and turnover in neural precursors. [25] CDCA8, also found downregulated in LND cells, is a component of a chromosomal passenger complex required for stability of the bipolar mitotic spindle.…”

Clinical severity in Lesch–Nyhan disease: The role of residual enzyme and compensatory pathways

“…The enzyme assay was conducted in live fibroblasts as previously described [15–17]. This assay involves the incorporation of radiolabeled hypoxanthine or guanine into related nucleotides.…”

“…Purines were measured by high performance liquid chromatography with photodiode array ultraviolet detection (HPLC-UV) as previously described [16, 17]. Cells were grown in an atmosphere of 5% CO 2 in DMEM, supplemented with 1 μg/mL of streptomycin, 1 units/mL of penicillin, 2 mM glutamine and 15% fetal calf serum (Invitrogen, Carlsbad, CA).…”

Is cIMP a second messenger with functions opposite to those of cGMP?

“…A straightforward explanation for this negative finding is that cellular ITP concentrations are very low so that there is no ITP substrate for nucleotidyl cyclase to act upon. In accordance with this hypothesis, sophisticated HPLC-MS/MS analysis of neuronal cells revealed that only inosine 5′-monophosphate (IMP) is present at low levels, but no ITP (Göttle et al 2013). The most important review in the field of cellular purine nucleotide concentrations (Traut 1994) also mentions just IMP as physiological cell constituent.…”