Glial cells produce myelin and contribute to axonal morphology in the nervous system. Two myelin membrane proteolipids, PLP and DM20, were shown to be essential for the integrity of myelinated axons. In the absence of PLP-DM20, mice assembled compact myelin sheaths but subsequently developed widespread axonal swellings and degeneration, associated predominantly with small-caliber nerve fibers. Similar swellings were absent in dysmyelinated shiverer mice, which lack myelin basic protein (MBP), but recurred in MBP*PLP double mutants. Thus, fiber degeneration, which was probably secondary to impaired axonal transport, could indicate that myelinated axons require local oligodendroglial support.

The tet regulatory system in which doxycycline (dox) acts as an inducer of specifically engineered RNA polymerase II promoters was transferred into transgenic mice. Tight control and a broad range of regulation spanning up to five orders of magnitude were monitored dependent on the dox concentration in the water supply of the animals. Administration of dox rapidly induces the synthesis of the indicator enzyme luciferase whose activity rises over several orders of magnitude within the first 4 h in some organs. Induction is complete after 24 h in most organs analyzed. A comparable regulatory potential was revealed with the tet regulatory system where dox prevents transcription activation. Directing the synthesis of the tetracycline-controlled transactivator (tTA) to the liver led to highly specific regula- A "genetic switch" that could be operated at will and that would permit the control of individual gene activities quantitatively and reversibly in a temporal and spatial manner would thus be of great advantage. The tetracycline (Tc)-controlled systems for the activation of transcription (7, 8) fulfill a number of these requirements at the cellular level. Herein, we report that the "reverse Tc-controlled transactivator" (rtTA) system, where doxycycline (dox) acts as an inducer of transcription as well as the "Tc-controlled transactivator" (tTA) system, where Tc or dox prevent transcription activation (Fig. 1) can be operated in a quantitative and highly tissue-specific way when transferred into mice. The results show that the controls are tight and that the kinetics of induction, especially with the rtTA system, are rapid. Although we (9, 10) and others (11,12) have reported that the tTA system can be applied to transgenic organisms, the results reported herein establish that both the rtTA and the tTA systems provide true genetic switches capable of quantitatively controlling individual gene activities in animals in a highly tissue-specific manner. These observations open up exciting prospects for the study of gene function in mammalian organisms. Transgenic Animals. Transgenic mouse lines (NMRI outbred) were generated by pronuclear injection using standard techniques (14) and analyzed by the Southern blot technique (15) using the BamHI-EcoRV fragment of the luciferase gene and the XbaI fragment of the tTA gene as respective probes. Luciferase reporter mice were obtained upon transfer of the 3.1-kb XhoI-EaeI fragment of pUHC13-3 (7). Mice producing rtTA controlled by PhCMV, the promoter of the human cytomegalovirus immediate early genes, were obtained upon transfer of the 2.7-kb PflMI-XhoI fragment and animals synthesizing tTA under the control of the LAP promoter were obtained by transferring the 5.5-kbAseI-Asp700 fragment of pUHG15-30. Animals transgenic for both a transactivator and a reporter unit were exposed when necessary to doxycycline hydrochloride (Sigma) dissolved in 5% sucrose supplied as drinking water, which was exchanged every 3 days. Possible long-term effects of dox (200 ,ug/ml) were exam...

Two proteolipid proteins, PLP and DM20, are the major membrane components of central nervous system (CNS) myelin. Mutations of the X-linked PLP/DM20 gene cause dysmyelination in mouse and man and result in significant mortality. Here we show that mutant mice that lack expression of a targeted PLP gene fail to exhibit the known dysmyelinated phenotype. Unable to encode PLP/DM20 or PLP-related polypeptides, oligodendrocytes are still competent to myelinate CNS axons of all calibers and to assemble compacted myelin sheaths. Ultrastructurally, however, the electron-dense 'intraperiod' lines in myelin remain condensed, correlating with its reduced physical stability. This suggests that after myelin compaction, PLP forms a stabilizing membrane junction, similar to a "zipper." Dysmyelination and oligodendrocyte death emerge as an epiphenomenon of other PLP mutations and have been uncoupled in the PLP null allele from the risk of premature myelin breakdown.