The gene Tousled of Arabidopsis Thaliana encodes a protein kinase which, when mutated, results in abnormal flower development. From a library of mRNAs that are translationally upregulated by overexpression of the translation initiation factor 4E, we identified a mammalian Tousled Like kinase (TLK1B). The human TLK1B mRNA contains a 5'UTR 1088-nt-long with two upstream AUG codons, and was found to be very inhibitory for translation. The TLK1B protein localizes almost exclusively to the nuclei. TLK1B overexpression in mammalian cells rendered them more resistant to ionizing radiation (IR). Purified TLK1B phosphorylated histone H3 at S(10) with high specificity both in a mix of core histones and in isolated chromatin, suggesting that histone H3 is a physiological substrate for TLK1B. Moreover, overexpression of TLK1B in transfected cells resulted in a higher degree of H3 phosphorylation. Expression of TLK1B in a yeast strain that harbors a temperature-sensitive mutation of the major H3 kinase, Ipl1, complemented the growth defect; restored normal levels of histone H3 phosphorylation; and increased their resistance to IR. Phosphorylation of H3 has been linked to the activation of the immediate-early genes upon mitogenic stimulation, and to chromatin condensation during mitotic/meiotic events. A possible role for TLK1B in radioprotection is discussed.