STOCK Tg(SFTPC-TK*)1Dris/J

Stock number: 032157
Product name: SPCTK transgenic mice (SPC-SR39TK)
Research areas: Apoptosis Research, Cell Biology Research, Developmental Biology Research, Internal/Organ Research, Research Tools

Description

SPCTK transgenic mice (SPC-SR39TK) allow gancyclovir (GCV)-inducible/reversible ablation of alveolar epithelial type 2 cells (AEC2). This is a model for studying AEC2 depletion in lung injury, repair and regeneration.

Detailed description

Type 2 alveolar epithelial cells (AEC2) are regarded as the progenitor population of the alveolus responsible for injury repair and homeostatic maintenance in distal lung.

The SPCTK transgene (SPC-SR39TK) has expression of the highly-sensitive SR39TK mutant Herpes simplex thymidine kinase 1 (HSV1-SR39TK) directed to AEC2 by the human surfactant protein C promoter (SPC). Because dividing/proliferating cells that express HSV1-TK will metabolize nucleoside analogs into toxic metabolites and undergo cell death, these SPCTK transgenic mice allow for temporally-regulated ablation of dividing AEC2 by systemic administration of the nucleoside analog ganciclovir (GCV). In addition, ablation of AEC2 is reversible upon GCV removal.

The phenotype of C57BL/6 SPCTK transgenic mice (hemizygous) is described below.
The high GCV sensitivity of SR39TK allows a wide dosable range with a 10-fold difference between minimal effective dose and lethal dose (10 and 100 mg/kg GCV intraperitoneal injection, respectively) - inducing specific and quantifiable levels of AEC2 depletion. Specifically, SPCTK mice treated with 10 mg/kg GCV caused only a modest AEC2 reduction (50%) and featured a significant rebound in AEC2 numbers over the course of recovery. In contrast, 100 mg/kg GCV treatment caused a significant AEC2 reduction (82%) that proved lethal at 13 days due to pulmonary edema. Of note, while a 50 mg/kg GCV treatment caused 80% AEC2 depletion over a slightly longer period (14 days), the majority of this cohort did not develop a fatal pulmonary phenotype (7% lethality). [Garcia et al. 2016 Am J Respir Cell Mol Biol. 54:319]
Analysis of several SPCTK-derived cell populations show GCV-directed SR39TK-induced killing in the lung is confined to SPC-expressing cells (AEC2). Active SR39TK was not present in SPCTK-derived fibroblasts or immune cells, and these populations were insensitive to GCV treatment. [Garcia et al. 2016 Am J Respir Cell Mol Biol. 54:319]

Prior to GCV exposure, these C57BL/6 SPCTK transgenic mice (both hemizygotes and homozygotes) are viable and fertile, with no reported gross phenotype, behavioral abnormalities or apparent cell death. Although the transgene is expressed in both male and female germlines, both male and female mice are fertile. However for homozygotes, the increased expression of the transgene is such that modulating GCV dosage is very difficult. Because the homozygous mice are extremely sensitive and AEC2 are lethally ablated at low GCV doses, the donating investigator routinely uses hemizygous mice for their experiments. In the absence of GCV injection, the donating investigator reports no apparent leakiness of the transgene - AEC2 levels in SPCTK and C57BL/6J mice do not significantly differ.

To date (July 2019), the donating investigator has not tested the efficacy of other nucleoside analogs - acyclovir (ACV) or the prodrug valganciclovir (VGCV) - in these C57BL/6 SPCTK transgenic mice.

This SPCTK transgene has some HIV-1 lentiviral genome sequences flanking the SPC-TK region. Importantly, the donating investigator reports that no infectious particles can be created from the HIV-1 genome sequences in this SPCTK transgene.

Development

SPCTK transgenic mice (SPC-SR39TK) were created by Dr. Barbara Driscoll (Children's Hospital Los Angeles, University of Southern California). Specifically, the donating investigator reports the 11069 bp construct pCCL-c-SPC-SR39TK was used for the SPCTK transgene. It contains (from 5' to 3') an ampicillin resistance cassette, a replication origin [ORI], a minimal human cytomegalovirus promoter/enhancer sequence, some HIV-1 lentiviral sequences (R region, truncated 5' LTR [U5], Ψ [packaging signal], reverse response element [RRE], central polypurine tract and central termination sequence [cPPT/CTS]), a 3.7 kbp human surfactant associated protein C promoter sequence (SFTPC; SPC), a Herpes simplex thymidine kinase 1 mutant SR39TK sequence (HSV1-SR39TK; with increased GCV prodrug catalytic activity) and additional HIV-1 lentiviral sequences (3' LTR, R region, truncated 5' LTR [U5]). Importantly, the donating investigator reports that no infectious particles can be created from the HIV-1 genome sequences in this SPCTK transgene.

The 11069 bp lentiviral vector was microinjected into C57BL/6J embryos. Specifically, this was by transducing ova with a non-replicating lentivirus packaged in a VSV capsid, then fertilizing the transduced ova in vitro. Founder male mice were bred to C57BL/6NCrl inbred mice for germline transmission. SPCTK transgenic mice from a single founder line were characterized further by PCR analysis and functional assessment following GCV injection. SPCTK transgenic mice were then bred to C57BL/6J inbred mice and/or C57BL/6-congenic Rosa26-LSL-lacZ mice (derived from Stock No. 003474) and/or C57BL/6N-congenic Sftpc-EGFP BAC transgenic mice (derived from Stock No. 028356). Next, the SPCTK colony was bred to C57BL/6J inbred mice for 5-6 generations (to remove Rosa26-LSL-lacZ and Sftpc-EGFP). In 2021, SPCTK males were sent to The Jackson Laboratory Repository. Upon arrival, sperm was cryopreserved. To generate the living colony, an aliquot of the frozen sperm was used to fertilize C57BL/6J oocytes (Stock No. 000664). The SPCTK colony was then maintained by the donating laboratory by breeding transgenic mice together, to wildtype (noncarrier) mice from the colony or to C57BL/6J inbred mice (see SNP note below).

In 2021, a 48 SNP (single nucleotide polymorphism) panel analysis, with 43 markers covering all 19 chromosomes and the X chromosome, as well as 5 markers that distinguish between the C57BL/6J and C57BL/6N substrains, was performed on the males sent to The Jackson Laboratory Repository. Five of the 43 markers throughout the genome were segregating with an unknown source. Also, 4 of the 5 markers that determine C57BL/6J from C57BL/6N were found to be segregating, including the rd8 recessive mutation associated with retinal degeneration on Chromosome 1 within the Crb1 locus. These data suggest the mice sent to The Jackson Laboratory Repository were on a STOCK genetic background.

Of note, the donating investigator reports that, at least once in this pedigree, an SPCTK female was bred to a C57BL/6J inbred male (thus the Y chromosome is expected to be C57BL/6J origin). The transgene insertion site(s) and transgene copy number have not yet been determined (July 2019). The donating investigator reports the transgene should not be able to jump around to a new location every generation (but did not test this specifically).

Allele

Symbol: Tg(SFTPC-TK*)1Dris
Name: transgene insertion 1, Barbara Driscoll
MGI accession ID: MGI:6331067
Generation method: Transgenic
Attributes: Inserted expressed sequence
Synonyms: SPCTK
Marker symbol: Tg(SFTPC-TK*)1Dris
Marker name: transgene insertion 1, Barbara Driscoll
Chromosome: UN
Strain of origin: C57BL/6J
Site of expression: The SPCTK transgene (SPC-SR39TK) has expression of the highly-sensitive SR39TK mutant Herpes simplex thymidine kinase 1 (HSV1-SR39TK) directed to type 2 alveolar epithelial cells (AEC2) by the human surfactant protein C promoter (SPC).
Molecular note: The transgene contains (from 5' to 3') an ampicillin resistance cassette, a replication origin, a minimal human cytomegalovirus promoter/enhancer sequence, some HIV-1 lentiviral sequences (R region, truncated 5' LTR [U5], psi [packaging signal], reverse response element, central polypurine tract and central termination sequence, a 3.7 kbp human surfactant associated protein C promoter sequence, a Herpes simplex thymidine kinase 1 mutant SR39TK sequence and additional HIV-1 lentiviral sequences (3' LTR, R region, truncated 5' LTR [U5]).