Cell death
Chinook salmon from the 18° treatment show a slight (two-fold to three-fold) increase in apoptosis-related gene expression, which increases up to seven-fold in the 21° treatment group. The 25° treatment group demonstrates a large increase in expression and number of upregulated apoptosis-related genes, ranging from two-fold to 220-fold change. Twelve out of 33 individuals in the 25° group became unresponsive and/or died during the 3-hr exposure. These individuals were monitored closely and removed for sampling promptly after cessation of opercular beats to avoid degradation of RNA in the sampled tissues.
At least 18 pro-apoptotic genes were upregulated by Chinook salmon following thermal exposure. The upregulation of CASP8, CASP10, RYBP, and KIAA0141 in the 25° group indicates that the intrinsic apoptotic pathway is active. Caspases 8 and 10 both participate in apoptotic cascades as well as increase NF-κb-mediated inflammation (Wang et al. 2001; Takahashi et al. 2005; Sakata et al. 2007). KIAA0141, death-associated protein effector DELE, is a pro-apoptotic protein that functions in the same pathway as caspases 8 and 10, and is functionally enhanced by inflammatory proteins such as TNFα (Harada et al. 2010). RYBP codes for death effector domain-associated factor, which binds with the protein hippy to increase the apoptotic effect of caspase 8 (Stanton et al. 2007). The apoptotic and inflammatory responses may be tightly intertwined during the response to stress observed in juvenile Chinook salmon. PPP1R13B, JUNB, ODC1 code for apoptosis-inducing proteins and were upregulated at 21° and 25° (Jacobs-Helber et al. 1998; Samuels-Lev et al. 2001; Bergamaschi et al. 2004). Interestingly, increases in ODC1 have been observed in moribund temperature-stressed adult salmonids (Pignatti et al. 2004), and it has functional roles beyond apoptosis, including involvement in cell proliferation.
There is some overlap between the UPR and induction of apoptosis (Didelot et al. 2006). CHAC1 (upregulated at 18°, 21°, and 25°) is a pro-apoptotic gene that is upregulated in response to the UPR (Mungrue et al. 2008). BAG3 (upregulated at 21° and 25°) is a regulator of many biological processes, among them apoptosis, and is a member of the BCL2 co-chaperone family that interacts directly with HSP70 (Rosati et al. 2011). Due to the multiple roles of these genes, it is uncertain which ultimate function they would serve in temperature-stressed fish in this experiment.
The processes of apoptosis and autophagy are tightly linked. Apoptosis results in cell death (type I), whereas autophagy can aid a cell in preventing apoptosis or can result in autophagic death (type II). Autophagy and apoptosis are mutually inhibitory to some extent, and both processes may be competing in a stressed cell (Mauiri et al. 2007). Given the mutual inhibition of autophagy and apoptosis, it is not surprising that we found upregulation of several genes that code for anti-apoptotic proteins, in addition to the pro-apoptotic genes discussed above. Genes in the BCL2 family regulate apoptosis and can be either pro- or anti-apoptotic. BCL2L1, BNIP3, MCL1, and BCL2L10 are all generally anti-apoptotic members of this family and were upregulated in the 25° treatment group (Lee et al. 0.1999; Desagher and Martinou 2000; Vande Velde et al. 2000; Sevilla et al. 2001; Craig 2002). Upregulated PIM1, another regulator of apoptosis, is usually anti-apoptotic, but it has also been shown to be pro-apoptotic in other circumstances (Lilly et al. 1999; Mochizuki et al. 1997; Aho et al. 2004; Gu et al. 2009).