Reply to “TPC1 Knockout Knocks Out TPC1”

REPLY

In a recent study (1), the pattern and intensity of proteins labeled with a photoaffinity probe (5N₃-[³²P]NAADP) were unchanged between samples from wild-type (WT) Tpcn1^LEXKO-471 mice, from which the authors concluded that TPC1 was not an NAADP-binding protein. However, no data were provided to assess the status of TPC1 expression in the Tpcn1^LEXKO-471 mutant mice.

In our recent study on a different Tpcn1 mutant mouse line (Tpcn1^T159) (generated by a gene trap insertion in the same Tpcn1 intron and in very close proximity to the insertion site of the gene trap in Tpcn1^LEXKO-471), we provided evidence that this gene trap locus did not significantly affect the expression of either Tpcn1A or Tpcn1B transcripts (2). Consequently, we expressed our concerns regarding the suitability of the similar Tpcn1^LEXKO-471 mice as a valid TPC1 knockout (KO) model and highlighting the need for a thorough characterization of the Tpcn1^LEXKO-471 line.

In their letter, Hooper et al. (3) now provide some data related to the status of Tpcn1 expression in Tpcn1^LEXKO-471 mice. Their immunoblot images show a significant reduction in the intensity of the immunoreactive band believed to correspond to TPC1A, and this therefore is good evidence that the Tpcn1^LEXKO-471 mutation has an impact on TPC1 expression.

However, with immunoblotting, one cannot rule out the possibility of residual expression of TPC1A or TPC1B, which might be unmasked by higher exposure and/or higher antibody concentration. Therefore, although the data presented by the authors go some way to characterize the Tpcn1 expression status in Tpcn1^LEXKO-471 mice, additional, more sensitive and definitive assessments of expression (e.g., reverse transcription-PCR data showing the lack of Tpcn1A and Tpcn1B transcripts) as well as functional assessments will be important to fully establish whether Tpcn1^LEXKO-471 is a complete TPC1 KO rather than a TPC1 hypomorphic model.