Puma and Trail/Dr5 pathways control radiation-induced apoptosis in distinct populations of testicular progenitors.

Abstract

Spermatogonia- stem cells and progenitors of adult spermatogenesis- are killed through a p53-regulated apoptotic process after gamma-irradiation but the death effectors are still poorly characterized. Our data demonstrate that both intrinsic and extrinsic apoptotic pathways are involved, and especially that spermatogonia can be split into two main populations, according to apoptotic effectors. Following irradiation both Dr5 and Puma genes are upregulated in the alpha6-integrin-positive Side Population (SP) fraction, which is highly enriched in spermatogonia. Flow cytometric analysis confirms an increased number of Dr5-expressing SP cells, and Puma-beta isoform accumulates in alpha6-integrin positive cellular extracts, enriched in spermatogonia. Trail-/- or Puma-/- spermatogonia display a reduced sensitivity to radiation-induced apoptosis. The TUNEL kinetics strongly suggest that the extrinsic and intrinsic pathways, via Trail/Dr5 and Puma respectively, could be engaged in distinct subpopulations of spermatogonia. Indeed flow cytometric studies show that Dr5 receptor is constitutively present on more than half of the undifferentiated progenitors (Kit- alpha6+ SP) and half of the differentiated ones (Kit+ alpha6+ SP). In addition after irradiation, Puma is not detected in the Dr5-positive cellular fraction isolated by immunomagnetic purification, while Puma is present in the Dr5-negative cell extracts. In conclusion, adult testicular progenitors are divided into distinct sub-populations by apoptotic effectors, independently of progenitor types (immature Kit-negative versus mature Kit-positive), underscoring differential radiosensitivities characterizing the stem cell/progenitors compartment.