Sequence organization and molecular cloning of mouse mammary tumor virus DNA endogenous to C57BL/6 mice

Abstract

The sequence organization of mouse mammary tumor virus DNA endogenous to the C57BL/6 inbred mouse strain was characterized by Southern blot analysis, utilizing probes specific for particular regions of the mouse mammary tumor virus provirus and by molecular cloning of endogenous mouse mammary tumor virus DNA. The genome of C57BL/6 mice contains three apparently intact, endogenous proviral units; two of these units comprise the Mtv-8 (unit II) and Mtv-9 (unit III) genetic loci that are also present in the DNA of BALB/c mice. The third unit is defined by EcoRI restriction fragments of 10.0 and 8.4 kilobases that contain the 5' and 3' portions of the provirus, respectively. This unit, termed unit XI and encoded by the genetic locus Mtv-17, has not been previously recognized in C57BL/6 DNA, but it can be clearly distinguished from the proviral units at Mtv-8 and Mtv-9 by Southern blot analysis under appropriate conditions. The proviral unit at Mtv-17 is not present in BALB/c DNA. DNAs comprising the entire Mtv-8 locus and the 3' portions of Mtv-9 and Mtv-17 were cloned. Analysis of the cloned DNA revealed no obvious deletions or rearrangements that would render proviral DNA defective; however, these endogenous genes are normally not transcriptionally active.

Insertion elements and transitions in cloned mouse mammary tumour virus DNA: further delineation of the poison sequences

“…The numbers involved are clearly too low to draw firm conclusions, but the distinction between exogenous and endogenous sequences raises an important issue. Several laboratories, including ours, have repeatedly cloned the 5' end of the endogenous Mtv-8 locus in both lambda and plasmid vectors and with no evidence for rearrangements (8,9,(13)(14)(15)(16). In contrast, we have only once isolated an Mtv-9 5' end, despite screening several large libraries of BALB/c DNA, though again the derived plasmid clone was apparently intact (10).…”

“…Numerous independent reports have alluded to problems encountered in manipulating MMTV DNA clones, but it is evident that these difficulties may be manifest in different ways. Thus, some laboratories have observed deletions, here we describe insertions and transitions, while in other situations the requisite clones could not be detected at all (1)(2)(3)(4)(5)(6)(7)(8)(9)(10). Many of these differences could be explained by variations in procedures, for example the choice of vectors, the strain of MMTV, and the use of integrated versus non-integrated proviral DNA or amplified versus non-amplified libraries.…”

“…In early attempts to clone the DNA provirus of mouse mammary tumour virus (MMTV), several laboratories reported either the anomalous behaviour or non-recovery of the required recombinants in otherwise representative libraries (1)(2)(3)(4)(5). These repeated observations led to the suggestion that certain MMTV sequences, for which the term "poison sequence" was coined, might be inimical to the growth of prokaryotic hosts, a concept that has been sustained by subsequent anecdotal reports and has until recently frustrated attempts to complete the characterisation of the MMTV genome (1)(2)(3)(4)(5)(6)(7)(8)(9)(10)(11).…”

“…In practice, particularly when dealing with milk-transmitted MMTV, the number of 3' junctions obtained far exceeds 5' junctions, and it was from this observation that the notion of a poison sequence was originally formed (1)(2)(3)(4)(5). Indeed, there have been only sporadic successes in the isolation of 5' junction fragments, and most of those have involved endogenous MMTV proviruses (8,10,(12)(13)(14)(15)(16)(17). Curiously, in extensive analyses of the milk-transmitted MMTV proviruses characteristic of the BR6 strain of mice, we have reproducibly isolated 5' and 3' ends with roughly equal frequency, in apparent contradiction to earlier reports.…”

Altered mouse mammary tumor virus transcript synthesis in T-cell lymphoma cells

“…MTV-1 and MTV-2 are the only endogenous proviral loci known to induce mammary tumors (20). Of the three proviral genomes of CS7BL/6 mice (the parental strain of EL-4 cells), none carry the env BamHI site (29), suggesting that the amplified provirus in EL-4 cells might have originated from a horizontally transmitted exogenous infectious MMTV.…”