Characterization of the mouse strain carbon(3)hafnium/selenium germinally-transmitted mtv proviruses: Analysis of the transcriptional regulatory region and identification of mtv-1 as the specific provirus template of mammary tissue mtv RNA transcripts

Date of Award




Degree Name

Doctor of Philosophy (Ph.D.)


Microbiology and Immunology

First Committee Member

Robert J. Pauley - Committee Chair


The C3Hf/Se mouse strain contains three full-length endogenous mouse mammary tumor virus proviruses in germline DNA, designated mtv-1, mtv-8, and mtv-11. C3Hf/Se mtv germline proviruses were analyzed on the DNA level for transcriptional activity in mammary tissues. The 3$\sp\prime$ EcoR I fragment of mtv-1, the C3Hf/Se endogenous provirus, implicated in tumorigenesis, was cloned into lambda gt 10. The 4.5 kbp insert contained 2.9 kbp of 3$\sp\prime$ mtv structural gene sequences, the 1.3 kbp terminal repeat (LTR) and 0.3 kbp of host DNA. For sequence homology and diversity between the U3 region of mtv-1 LTR, mtv-8 U3 region, a 1.5 kbp Pst I-EcoR I fragment containing mtv-1 LTR and host sequences was subcloned into the M13 sequencing vectors. The 1.3 kbp LTR of mtv-1 was sequenced using synthetic oligonucleotides. No sequence modifications of the mtv-1 LTR explained its transcriptional activity. Other mtv proviruses remain cryptic. Characterization of mtv-1 LTR revealed sequence heterogeneity compared to the same region in mtv-8 LTR. Optimal sequence alignment of mtv-1 to mtv-8 LTR sequence revealed 123 bases in mtv-8 LTR was replaced with 116 bases within mtv-1 in which there were 73 unaligned bases compared to mtv-8, due to base mismatches, additions or deletions. This heterogeneity located 5$\sp\prime$ to the transcriptional regulatory sequences in mtv-1 altered the open reading frame (orf) of mtv-1 LTR. Computer translation of mtv-1 orf showed total disruption of COOH-terminus of putative orf proteins. Mtv-11 LTR transcriptional regulatory region was compared to mtv-8 LTR sequence. Promoter and regulatory regions were similar to mtv-8 and mtv-1 DNA sequence. The area 5$\sp\prime$ to the transcriptional regulatory sequences in mtv-11 U3 region unlike sequence alterations in mtv-1 LTR. Mtv-11 sequences of the LTR were 100% homologous to mtv-8 LTR sequences. The 116 base pair sequence alteration in the U3 region of mtv-1 LTR identified the mtv provirus template(s) for mtv RNA transcripts that accumulate in C3Hf/Se mammary tissues by S1 nuclease mapping. Results from the S1 nuclease mapping experiments demonstrated that mtv-1 is transcriptionally active in C3Hf/Se mammary tumors whereas mtv-8 and mtv-11 are either not transcriptionally active or are expressed at levels too low to be detected by the S1 method. (Abstract shortened with permission of author.)


Biology, Microbiology

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