Snm1-deficient mice exhibit accelerated tumorigenesis and susceptibility to infection

Abstract

The eukaryotic SNM1 gene family has been implicated in a number of cellular pathways, including repair of DNA interstrand cross-links, involvement in VDJ recombination, repair of DNA double-strand breaks, and participation in cell cycle checkpoint pathways. In particular, mammalian SNM1 has been shown to be required in a mitotic checkpoint that causes arrest of cells in prophase prior to chromosome condensation in response to spindle poisons. Here, we report on the phenotype of a knockout of Snm1 in the mouse. Snm1-/- mice are viable and fertile but exhibit a complex phenotype. Both homozygous and heterozygous mice show a decline in survival compared to wild-type littermates. In homozygous mutant males, this reduction in survival is principally due to bacterial infections in the preputial and mandibular glands and to a lesser extent to tumorigenesis, while in homozygous and heterozygous females, it is due almost solely to tumorigenesis. The high incidence of bacterial infections in the homozygous mutant males suggests an immune dysfunction; however, examinations of T- and B-cell development and immunoglobulin class switching did not reveal a defect in these pathways. Crossing of Snm1 mutant mice with a Trp53 null mutant resulted in an increase in mortality and a restriction of the tumor type to lymphomas, particularly those of the thymus. Taken together, these findings demonstrate that Snm1 is a tumor suppressor in mice that in addition has a role in immunity.

FIG. 1.

Disruption of Snm1 in the mouse by gene targeting and genotype of knockout mice. (A) Schematic representations of mouse Snm1 locus (top) and the Snm1 targeting vector (middle). Solid boxes denote exons, with 5′ ends to the left. D (DraI), B (BstZ171), E (EcoRI), and S (SstI), restriction enzyme sites used for analyses of the targeted deletion. Prob, probe. (B) Southern blot analysis of tail DNA from F₂ animals. The sizes of SstI fragments are indicated. Genomic DNA digested with SstI and hybridized with the 5′ probe yielded fragments of 4.3 and 3 kb in the wild-type and targeted locus, respectively (left). Hybridization with the 3′ probe yielded fragments of 12 and 6.4 kb in the wild-type and targeted locus, respectively (right). (C) Multiplex PCR genotype analysis with primer pair S1/S2 (lanes 1 and 2), which results in a 602-bp product in the wild-type allele, and with primer pair S1/N1 (lanes 2 and 3), which resulted in a 240-bp product in the targeted allele.

FIG. 2.

Survival of Snm1^−/− mice. (A) Increased mortality in Snm1 mice shown by Kaplan-Meier analysis. Snm1^−/−, Snm1^+/−, and Snm1^+/+ animals were observed for 24 months. (B) Kaplan-Meier analysis showing survival by gender and genotype of Snm1^−/− male, Snm1^+/− male, Snm1^+/+ male, Snm1^−/− female, Snm1^+/− female, and Snm1^+/+ female cohort mice versus age in months.

FIG. 3.

Incidence of infections in Snm1 mice. (A) Frequencies of bacterial infections in preputial and mandibular glands of Snm1 mice up to 24 months of age. (B) Examples (arrows) of infected organs, mandibular gland (upper) and preputial gland (lower), obtained from Snm1^−/− mice.

FIG. 4.

Effects of infections and tumorigenesis on the mortality of Snm1 mice. The number of mice within each genotype that were followed for tumor development and infections and the mean risk values are given in Tables 1 and 2, respectively. (A) Kaplan-Meier tumor-free-survival curve by gender and genotype of Snm1 cohort mice versus age in months. (B) Kaplan-Meier infection-free-survival curve by gender and genotype of Snm1 cohort mice versus age in months.

FIG. 5.

Histologic analysis of tumors found in Snm1^−/− mice. (a to c) Examples of spontaneous tumors from different organs of Snm1^−/− mice: (a) liver, (b) adrenal gland, and (c) uterus and ovary. (d to f) Representative histology of lymphomas of Snm1^−/− mice: (d) mandibular lymph node lymphoma, ×100 magnification; (e) spleen lymphoma, ×50 magnification; and (f) adrenal gland lymphoma, ×200 magnification. (g to i) Representative histology of adenomas of Snm1^−/− mice: (g) lung adenoma, ×20 magnification; (h) papillary hardarian gland adenoma, ×50 magnification; and (i) ovary cyst adenoma, ×200 magnification. All tumors were obtained from 15-month- to 20-month-old Snm1^−/− mice. The tissues were paraffin embedded and stained with hematoxylin and eosin stain.

FIG. 6.

Homozygosity for Snm1 decreases the survival of Trp53^−/− mice. Kaplan-Meier analysis showing the survival of Snm1^−/− Trp53^−/−, Snm1^+/+ Trp53^−/−, and Snm1^−/− Trp53^+/− cohort mice versus age in months.

FIG. 7.

Snm1 homozygous mice are hypersensitive to the spindle poison taxol. Kaplan-Meier analysis showing the survival of Snm1^−/− and Snm1^+/+ mice versus age after injection of taxol at 15 mg/kg of body weight.