In vivo Role of TGase 3 Figure 7. Morphological changes in pelage hair lacking TGase 3. Whole-mount light microscopy of TGM32/2 (A) and wild type (B) hair (scale bar 10 mm). Inserts show higher magnification of lower regions of the hairs. Hair incubated overnight at 65uC with agitation in 2% SDS and 20 mM dithiothreitol (C–F) TGM32/2 (C, E) and control hairs (D, F). Arrowheads indicate regions lacking or retaining the cuticle cells in the mutant and control hairs, respectively. Scanning electron microscopy of mutant (G, I) and control (H, J) hair shafts shows severe distortion of the hair, in particular the cuticle, in the absence of TGase 3. In contrast, the roots of the mutant hairs (K) appear similar to the wild type ones (L) (scale bar 20 mm). Each of the four main pelage hair types was present in the mutant animals (M). doi:10.1371/journal.pone.0034252.g007 follicle degeneration. Interestingly, as in TGM32/2 mice, the phenotype improves after the first postnatal hair cycle. This finding and the genetic strain specificity appears to be caused by a compensatory increase in keratin16 levels [45]. Curiously, in both the K172/2 and TGM32/2 mice there is no nail defect despite both genes being expressed in the nail bed, even though missense mutations in keratin17 are related to type 2 pachyonychia congenita, which presents with severe nail dystrophy [44] and pili torti [46]. Though severe twisting in the hair shaft, present in human pili torti, was generally not observed in these mice, some hairs did show constrictions and torsions. Other human recessive forms of pili torti which improve with age have been described [47] and it is possible that these are related to changes in TGase 3 or its substrates. Indeed, recent findings have shown that variation in the trichohyalin gene associates with the straight hair phenotype in man [48]. Humans with lamellar ichthyosis have severe hair abnormalities [49] and skingrafted from murine TGM12/2 neonates develop abnormal hair follicles [50]. However, it is not known whether this is caused by specific changes in hair development or by the overall disruption of the grafted tissue. It would appear that TGase 1 is crucial for the development of the normal hair follicle, while TGase 3 plays a more subtle role, particularly in the formation of the hair cuticle. As in the skin, TGase 3 expression occurs later in hair maturation than TGase 1 [24], and it may act to reinforce the developing cornified envelope initiated by TGase 1, by introducing trichohyalin and keratin17/ keratin6hf containing intermediate filaments into the cornified envelope. There is much circumstantial evidence of a role for transglutaminases in wound healing. Surprisingly, there was little change in the rate of wound closure upon the loss of TGase 3. In immature wounds, keratinocytes express a wide spectrum of keratins, which become limited as the tissue matures. Post PLoS ONE | www.plosone.org 8 April 2012 | Volume 7 | Issue 4 | e34252