Characterization of transgenic mice with overexpression of spermidine synthase

Abstract

A composite cytomegalovirus-immediate early gene enhancer/chicken β-actin promoter (CAG) was utilized to generate transgenic mice that overexpress human spermidine synthase (SpdS) in order to determine the impact of elevated spermidine synthase activity on murine development and physiology. CAG-SpdS mice were viable and fertile and tissue SpdS activity was increased up to 9-fold. This increased SpdS activity did not result in a dramatic elevation of spermidine or spermine levels but did lead to a 1.5 to 2-fold reduction in tissue spermine:spermidine ratio in heart, muscle and liver tissues with the highest levels of SpdS activity. This new mouse model enabled simultaneous overexpression of SpdS and other polyamine biosynthetic enzymes by combining transgenic animals. The combined overexpression of both SpdS and spermine synthase (SpmS) in CAG-SpdS/CAG-SpmS bitransgenic mice did not impair viability or lead to overt developmental abnormalities but instead normalized the elevated tissue spermine:spermidine ratios of CAG-SpmS mice. The CAG-SpdS mice were bred to MHC-AdoMetDC mice with a >100-fold increase in cardiac S-adenosylmethionine decarboxylase (AdoMetDC) activity to determine if elevated dcAdoMet would facilitate greater spermidine accumulation in mice with SpdS overexpression. CAG-SpdS/MHC-AdoMetDC bitransgenic animals were produced at the expected frequency and exhibited cardiac polyamine levels comparable to MHC-AdoMetDC littermates. Taken together these results indicate that SpdS levels are not rate limiting in vivo for polyamine biosynthesis and are unlikely to exert significant regulatory effects on cellular polyamine content and function.

The function of spermine

“…This provides a great deal of discrimination in favor of the smaller substrate (more than 100‐fold). However spermidine synthase is present in much larger amounts than SpmSyn [in mice approximately 25 times in heart; 400 times in liver, and 40 times in brain ] Thus, even 0.5% activity would provide a considerable capacity to produce spermine, particularly, since levels of dcAdoMet, the other substrate for the reaction, are increased.…”

“…However, several altered residues could limit this space even more either directly or by slightly changing the position of Trp28. Transgenic mice in which human spermidine synthase is expressed from a ubiquitous promoter are available and cells derived from crosses of these mice with the Gy mice could be used to test this hypothesis.…”

Hybrid expression cassettes consisting of a milk protein promoter and a cytomegalovirus enhancer significantly increase mammary‐specific expression of human lactoferrin in transgenic mice

“…In summary, our ternary hybrid promoter/enhancer design containing a casein or a BLG promoter, a CMV enhancer, and a binary chicken β‐globin insulator can be considered a first step toward creating and validating an improved mammary gland‐specific vector, which ensures high‐level expression of the transgene, independent of the integration site. This newly developed vector may be used as a robust promoter for gene‐specific expression, particularly to direct mammary‐specific expression of transgenes, similar to the CAG promoter that consists of the CMV enhancer, the chicken β‐actin promoter, and an intron (Xu et al, 2001; Ito et al, 2006; Shi et al, 2012). Further confirmation of the efficacy of this unique hybrid promoter/enhancer vector is currently underway in domestic animals, such as rabbit, goat, pig, and cattle.…”

Expression of polyamine metabolism-related genes during the development of prehierarchical follicles in the Zi geese (Ansercygnoides)

“…The speculated variation in these results can be associated to the breed specific difference complex. Mostly abundant polyamines in the animal cells include spermidine, which is produced through the conversion of putrescine by spermidine synthase (SRM) in the polyamine biosynthesis pathway to promote cell growth (Shi et al 2012) . The results showed that the SRM gene was highly expressed in PE, with minimal expression in MWF, in which there was no remarkable expression difference between the SWF and SYF (p > 0.05) but significantly lower than LWF (p  0.05).…”