Fetal echocardiography

Summary available; p. 7-10

Timed degradation of Mcl-1 controls mitotic cell death

Mitotic arrest can result in cell death through the process of apoptosis. We have shown by live-cell imaging that the ubiquitin-proteasome dependent proteolysis of the apoptotic regulator Mcl-1 under the control of the anaphase-promoting complex or cyclosome (APC/C) provides a timing mechanism that distinguishes prolonged mitotic arrest from normal mitosis

USP9X limits mitotic checkpoint complex turnover to strengthen the spindle assembly checkpoint and guard against chromosomal instability

Faithful chromosome segregation during mitosis depends on the spindle assembly checkpoint (SAC), which delays progression through mitosis until every chromosome has stably attached to spindle microtubules via the kinetochore. We show here that the deubiquitinase USP9X strengthens the SAC by antagonizing the turnover of the mitotic checkpoint complex produced at unattached kinetochores. USP9X thereby opposes activation of anaphase-promoting complex/cyclosome (APC/C) and specifically inhibits the mitotic degradation of SAC-controlled APC/C substrates. We demonstrate that depletion or loss of USP9X reduces the effectiveness of the SAC, elevates chromosome segregation defects, and enhances chromosomal instability (CIN). These findings provide a rationale to explain why loss of USP9X could be either pro- or anti-tumorigenic depending on the existing level of CIN. Skowyra et al. show the deubiquitinase USP9X limits activation of the ubiquitin ligase APC/C during mitosis. Loss of USP9X causes chromosomal instability (CIN), which can promote cancer. This work also provides a rationale for targeting USP9X when it is expressed in cancer cells with high levels of CIN

Atypical APC/C-dependent degradation of Mcl-1 provides an apoptotic timer during mitotic arrest

The initiation of apoptosis in response to the disruption of mitosis provides surveillance against chromosome instability. Here, we show that proteolytic destruction of the key regulator Mcl-1 during an extended mitosis requires the anaphase-promoting complex or cyclosome (APC/C) and is independent of another ubiquitin E3 ligase, SCF Using live-cell imaging, we show that the loss of Mcl-1 during mitosis is dependent on a D box motif found in other APC/C substrates, while an isoleucine-arginine (IR) C-terminal tail regulates the manner in which Mcl-1 engages with the APC/C, converting Mcl-1 from a Cdc20-dependent and checkpoint-controlled substrate to one that is degraded independently of checkpoint strength. This mechanism ensures a relatively slow but steady rate of Mcl-1 degradation during mitosis and avoids its catastrophic destruction when the mitotic checkpoint is satisfied, providing an apoptotic timer that can distinguish a prolonged mitotic delay from normal mitosis. Importantly, we also show that inhibition of Cdc20 promotes mitotic cell death more effectively than loss of APC/C activity through differential effects on Mcl-1 degradation, providing an improved strategy to kill cancer cells

Xenopus tropicalis allurin: Expression, purification, and characterization of a sperm chemoattractant that exhibits cross-species activity

AbstractPreviously we reported the identification of the first vertebrate sperm chemoattractant, allurin, in the frog Xenopus laevis (Xl) and demonstrated that it was a member of the CRISP family of proteins. Here we report identification, purification, and characterization of Xenopus tropicalis (Xt) allurin, a homologous protein in X. tropicalis. “Egg water” as well as purified allurin from both species exhibit efficient cross-species sperm chemoattractant activity. Western blots show that Xt egg water contains a single anti-allurin cross-reactive protein whose molecular weight (20,497 Da by MALDI MS) agrees well with the molecular weight of the hypothetical gene product for a newly recognized “Crisp A” gene in the X. tropicalis genome. A recombinant form of the protein, expressed in 3T3 cells, exhibits chemoattraction for both Xt and Xl sperm and cross reacts with anti-allurin antibodies. Examination of Crisp protein expression in the Xt oviduct using RT-PCR showed that of five documented Xt Crisp genes (Crisps 2, 3, LD1, LD2 and A) only Crisp A was expressed. In contrast, Crisp 2, Crisp 3, Crisp LD1, and Crisp LD2, but not Crisp A, were all found to be expressed in the Xt testes while subsets of Crisp proteins where expressed in the Xt ovary. These data suggest that Crisp proteins in amphibians may play multiple roles in sperm production, maturation and guidance just as they are thought to in mammals indicating that Crisp protein involvement in reproduction may not be limited to mammals