Human CDK10 gene isoforms.

The CDK10/PISSLRE gene has been shown to encode two different CDK-like putative kinases. The function(s) of the gene products are unknown, although a role at the G2/M transition has been suggested. We characterised two novel cDNAs. CDK10 mRNA quantity was not found to be correlated with cell proliferation status in HeLa or WI38 cell cultures or in human tissues. Relative levels of the four CDK10 isoforms were studied by RT-PCR, of which three were principally expressed. The two initially cloned isoforms predominated in human tissues, except in brain and muscle. Relative isoform levels did not vary during the cell cycle in culture, except when cells entered into the cell cycle. Finally, the predominant isoforms were shown to have different translation initiation sites and to have different subcellular distribution, due to an alternatively spliced nuclear localisation signal.

Accelerated telomere shortening and telomerase activation in Fanconi's anaemia.

Fanconi's anaemia (FA) is an autosomal recessive disorder characterized by progressive bone marrow failure that often evolves towards acute leukaemia. FA also belongs to a group of chromosome instability diseases. Because telomeres are directly involved in chromosomal stability and in cell proliferation capacity, we examined telomere metabolism in peripheral blood mononuclear cells (PBMC). Telomere length was significantly shorter in 54 FA patient samples, compared to 51 controls (P<0.0001). In addition, mean telomere terminal restriction fragment lengths (TRF) in nine heterozygous patient samples did not differ from those of controls. In 14 samples from FA patients with severe aplastic anaemia (SFA), telomere length was significantly shorter than in 22 samples of age-matched FA patients with moderate haematological abnormalities (NSFA) (P<0.001). However, no correlation was found between TRF length and the presence of bone marrow clonal abnormalities in 16 additional, separately analysed, patient samples. Sequential measurement of TRF in six FA patients showed an accelerated rate of telomere shortening. Accordingly, telomere shortening rate was inversely correlated with clinical status. Telomerase, the enzyme that counteracts telomere shortening, was 4.8-fold more active in 25 FA patients than in 15 age-matched healthy controls. A model for the FA disease process is proposed.

Characterization of the components of reconstituted Saccharomyces cerevisiae RNA polymerase I transcription complexes.

We have reconstituted specific RNA polymerase I transcription from three partially purified chromatographic fractions (termed A, B, and C). Here, we present the chromatographic scheme and the initial biochemical characterization of these fractions. The A fraction contained the RNA polymerase I transcription factor(s), which was necessary and sufficient to form stable preinitiation complexes at the promoter. Of the three fractions, only fraction A contained a significant amount of the TATA binding factor. The B fraction contributed RNA polymerase I, and it contained an essential RNA polymerase I transcription factor that was specifically inactivated in response to a significant decrease in growth rate. The function of the C fraction remains unclear. This reconstituted transcription system provides a starting point for the biochemical dissection of the yeast RNA polymerase I transcription complex, thus allowing in vitro experiments designed to elucidate the molecular mechanisms controlling rRNA synthesis.