We report here for the first time that the cyclin-dependent kinase PITSLRE/CDK11 is a new factor regulating autophagy. We show that depletion of PITSLRE in Drosophila cells and CDK11, the one ortholog of PITSLRE in human cells, both result in accumulation of autophagosomes indicating that the modulation of autophagy by the kinase is evolutionarily conserved. We also show in human cells that the initial depletion of CDK11 causes cells to undergo an initial induction of autophagic flux whereas at later time points it is clear that autophagosome turnover is significantly retarded, although not completely blocked. How then can we explain these different effects on autophagy at different times? As no substrates of PITSLRE are currently known, it is only possible to speculate as to the role played by PITSLRE/CDK11 in autophagy and how it can have different effects at different times. One possibility is that at early time points the depletion of CDK11 results in induction of autophagy, but at later time points loss of CDK11 also results in a decrease in lysosomal function. This potential explanation is consistent with the data we provide showing that while at later time points autolysosomes are clearly formed, their turnover is markedly reduced ().
One interesting observation from our studies is the downregulation of p62
mRNA following protracted CDK11 knockdown (Fig. S2
). We can only assume that p62
mRNA levels decrease following CDK11 knockdown through a mechanism that decreases either p62
transcription or p62
mRNA stability. Previous studies have shown that p62 protein levels feed-forward to regulate p62
transcription via the transcription factor NRF2.23
Following prolonged CDK11 knockdown, p62 may become sequestered in autophagosomes/autolysosomes which are retarded in their turnover. As a result, this may well break the feed-forward loop regulating p62
transcription leading to decreased levels of p62
mRNA. Further studies are therefore required to ascertain if this mechanism is the cause of p62
mRNA downregulation following long-term loss of CDK11.
Although no clear mechanistic functions for CDK11 are known, in terms of targets for phosphorylation, the kinase has been reported to have effects on multiple biological functions including apoptosis, RNA splicing and mitosis.14–19
It is important therefore not to simply focus on how CDK11 regulates components of the autophagy machinery to bring about the effects we report here, but also how the effects of CDK11 on autophagy impinge on these other CDK11 functions and vice versa. In this regard, we examined the effects of CDK11 knockdown on cell cycle progression and cell death in MDA-MB-231 cells. This revealed that in the time frame studied, CDK11 knockdown had no effect on apoptosis as assessed by analysis for sub-G1
DNA content—a reliable marker of apoptotic death (Fig. S3
Moreover, accumulation of autophagosomes following CDK11 knockdown was not inhibited with a concentration of the pan-caspase inhibitor, zVAD-fmk that was sufficient to inhibit TNFα-driven apoptotic cell death (Fig. S4A and B
). A small increase (15%) in cells in G2
/M phase was observed following 72 h of CDK11 knockdown (Fig. S3
), but we do not consider this to be the cause of autophagy inhibition, since 70–80% of the cells exhibited marked autophagosome accumulation at this time point ().
CDK11 has been reported to be perturbed in various human cancers including neuroblastoma, non-Hodgkin's lymphoma and melanoma.25–27
In light of the findings we report here, it is natural to consider whether the regulation of autophagy is the reason for selection for alterations in CDK11 in these forms of cancer or again whether the involvement of CDK11 in other cellular functions is more important. Ultimately, extensive further studies are required to not only determine the role of CDK11 in the regulation of autophagy, but also to ascertain if CDK11 represents another control point at which autophagy can be deregulated in human disease.