Absence of signal peptide peptidase in peripheral sensory neurons affects latency-reactivation in HSV-1 ocularly infected mice
Fig 1
Development of sensory neuron specific SPP conditional knockout mice.
(A) Confirmation of Avil-SPP-/- moue genotype. Sensory neuron specific SPP knockout mice (Avil-SPP-/-) were generated by crossing SPPflox/flox mice with Advillin-Cre+ mice to generate Avil-SPP-/- mice, in which SPP was specifically knocked out in Advillin-expressing neuronal tissue. Mouse genotypes were confirmed by PCR as shown and mice 6 and 7 were confirmed to be Avil-SPP-/- (SPPflox/floxAvil-Cre+); (B) Lack of SPP expression was confirmed in TG of Avil-SPP-/- mice by IHC. TG of 2 month old Avil-SPP-/- mice or control mice were harvested and frozen in OCT compound at -80°C for further processing. IHC was performed with anti-SPP antibody and developed by VectaShield VIP substrate kit. SPP positive cells (purple color) were more abundant in TG from Avil-SPP-/- mice than from the control group; C) Quantitation of SPP expression density in TG of Avil-SPP-/- mice. SPP expression from eight separate IHC staining as (B) above were quantitated by image analysis; and (D) Lack of SPP expression in TG of Avil-SPP-/- mice confirmed by qRT-PCR. TG of 2-month-old Avil-SPP-/- or control mice were harvested, total individual TG RNA or RNA from isolated neurons were extracted, and qRT-PCR was performed. Levels of RNA expression in total TG RNA or neuronal RNA were normalized to that of RNA from their respective control mice. Fold change represents the mean ± SEM from six replications (6 TG for total RNA isolation and 24 TG for RNA isolation from neurons).