Chemical Characterization of Mainstream Smoke from SPECTRUM Variable Nicotine Research Cigarettes.

OBJECTIVE: Our objective was to characterize mainstream smoke constituent deliveries from SPECTRUM variable nicotine research cigarettes under 2 machine smoking regimens. SPECTRUM cigarettes are manufactured by the 22nd Century company for the National Institute on Drug Abuse, National Institutes of Health to contain varying (including reduced) levels of nicotine. METHODS: Mainstream smoke constituent deliveries of "tar," nicotine, carbon monoxide, tobacco-specific nitrosamines (N'-nitrosonornicotine (NNN) and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)), benzo[a]pyrene, aromatic amines, and carbonyls were analyzed in 23 varieties of SPECTRUM cigarettes using ISO 17025 accredited methods. RESULTS: Data are presented as means and standard deviations of 5 replicates for all analytes. CONCLUSIONS: Under the ISO smoking regimen, mean levels of many smoke emissions for SPECTRUM varieties were comparable to the 3R4F research cigarette. Calculated SPECTRUM elasticity ranged from 1.6 to 4.0. Accordingly, under intense machine smoking conditions differences in emissions of SPECTRUM cigarettes were apparent. In addition, NNN increased with smoke nicotine while the same rate of change was not seen for NNK. It is important to monitor levels of chemicals of public health concern and regulatory interest as technologies emerge to reduce levels of nicotine or other targeted chemicals in tobacco products.

The Ty1 Retrotransposon Restriction Factor p22 Targets Gag.

A novel form of copy number control (CNC) helps maintain a low number of Ty1 retrovirus-like transposons in the Saccharomyces genome. Ty1 produces an alternative transcript that encodes p22, a trans-dominant negative inhibitor of Ty1 retrotransposition whose sequence is identical to the C-terminal half of Gag. The level of p22 increases with copy number and inhibits normal Ty1 virus-like particle (VLP) assembly and maturation through interactions with full length Gag. A forward genetic screen for CNC-resistant (CNCR) mutations in Ty1 identified missense mutations in GAG that restore retrotransposition in the presence of p22. Some of these mutations map within a predicted UBN2 domain found throughout the Ty1/copia family of long terminal repeat retrotransposons, and others cluster within a central region of Gag that is referred to as the CNCR domain. We generated multiple alignments of yeast Ty1-like Gag proteins and found that some Gag proteins, including those of the related Ty2 elements, contain non-Ty1 residues at multiple CNCR sites. Interestingly, the Ty2-917 element is resistant to p22 and does not undergo a Ty1-like form of CNC. Substitutions conferring CNCR map within predicted helices in Ty1 Gag that overlap with conserved sequence in Ty1/copia, suggesting that p22 disturbs a central function of the capsid during VLP assembly. When hydrophobic residues within predicted helices in Gag are mutated, Gag level remains unaffected in most cases yet VLP assembly and maturation is abnormal. Gag CNCR mutations do not alter binding to p22 as determined by co-immunoprecipitation analyses, but instead, exclude p22 from Ty1 VLPs. These findings suggest that the CNCR alleles enhance retrotransposition in the presence of p22 by allowing productive Gag-Gag interactions during VLP assembly. Our work also expands the strategies used by retroviruses for developing resistance to Gag-like restriction factors to now include retrotransposons.