Novel and Recurrent Mutations in Keratin 1 Cause Epidermolytic Ichthyosis and Palmoplantar Keratoderma F

Concise report CED Clinical and Experimental Dermatology

Novel and recurrent mutations in keratin 1 cause epidermolytic ichthyosis and palmoplantar keratoderma F. J. D. Smith,1,2 I. M. Kreuser-Genis,3 C. S. Jury,4 N. J. Wilson,1 A. Terron-Kwiatowski5 and M. Zamiri6 1Dermatology and Genetic Medicine, Division of Biological Chemistry and Drug Discovery, School of Life Sciences, University of Dundee, Dundee, UK; 2Pachyonychia Congenita Project, Holladay, UT, USA; 3Department of Dermatology, University Hospital Crosshouse, Kilmarnock, UK; 4Department of Dermatology, Glasgow Royal Inﬁrmary, Glasgow, UK; 5East of Scotland Regional Genetics Service, Ninewells Hospital, Dundee, UK; and 6Alan Lyell Centre for Dermatology, Queen Elizabeth University Hospital, Glasgow, UK doi:10.1111/ced.13800

Summary Mutations in keratin genes underlie a variety of epidermal and nonepidermal cell- fragility disorders, and are the genetic basis of many inherited palmoplantar keratodermas (PPKs). Epidermolytic PPK (EPPK) is an autosomal dominant disorder that can be due to mutations in the keratin 1 gene, KRT1. Epidermolytic ichthyosis (EI), the major keratinopathic ichthyosis, is characterized by congenital erythroderma, blistering and erosions of the skin. Causative mutations in KRT1 and KRT10 have been described, with PPK being present primarily in association with the former. We report four unrelated cases (one with sporadic EI and three with autosomal dominant PPK), due to two novel and two recurrent KRT1 mutations. Mutations in KRT1 are not only scattered throughout the keratin 1 protein, as opposed to being clustered, but can result in a range of phenotypes as further conﬁrmed by these mutations, giving a complex genotype/phenotype pattern.

Keratins constitute the intermediate filament proteins have been described, with PPK being present primarily of keratinocytes, with the keratin cytoskeleton acting in association with the former. as an important structural stabilizer of epithelial cells.1 We report four unrelated cases with KRT1 muta- Mutations in keratin genes underlie a variety of epi- tions (one with sporadic EI and three with autosomal dermal and nonepidermal cell-fragility disorders, and dominant PPK) due to two previously unreported and are the genetic basis of a number of the inherited two reported KRT1 mutations. palmoplantar keratodermas (PPKs).2 Epidermolytic PPK (EPPK) (Vorner type; OMIM 144200) is an auto- Report somal dominant disorder that can be caused by dominant-negative mutations in the keratin 1 gene, KRT1.3 Genetic testing was performed following ethics Epidermolytic ichthyosis (EI) [OMIM 113800; also approval by an institutional review board. The study known as epidermolytic hyperkeratosis (EHK) and bul- complied with principles of the Declaration of Helsinki, lous congenital ichthyosiform erythroderma (BCIE)], and informed consent was obtained from all partici- the major keratinopathic ichthyosis, is characterized pants. by congenital erythroderma, blistering and erosions of Genomic DNA was extracted from peripheral blood the skin.4 Causative mutations in KRT1 and KRT10 leucocytes or from saliva collected using an Oragene DNA sample collection kit (DNA Genotek, Ontario, Correspondence: Dr Frances J. D. Smith, Pachyonychia Congenita Project, Canada). PCR and Sanger sequencing was performed PO box 17850, Holladay, UT, USA to screen the coding regions and exon/intron bound- E-mail: [email protected] aries of KRT1 (primer sequences available on request). Conflict of interest: the authors declare that they have no conflicts of Variants were confirmed as pathogenic by reference to interest. an in silico prediction tool (Mutation Taster; http:// Accepted for publication 30 July 2016 mutationtaster.org/) and by sequencing of other family

528 Clinical and Experimental Dermatology (2019) 44, pp528–534 ª 2018 British Association of Dermatologists Novel and recurrent mutations in keratin 1 F. J. D. Smith et al.

Family 1 (a) (b)

(c)

(d) Wild type K1, exon 1 (e) K1 p.Leu187Pro

_ Figure 1 Family 1: the proband had (a) K S L N N K S - N N hyperkeratosis of the digits and palm, (b) diffuse plantar hyperkeratosis and (c) warty ﬂexural hyperkeratosis. (d) Wild- type KRT1 sequence of exon 1 (nucleotides 553–567) and (e) the equivalent region of KRT1 from the proband. The arrow indicates a heterozygous mutation c.560T>C resulting in an amino acid substitution of leucine to proline, p.Leu187Pro, in the keratin 1 protein. members when available. None of the variants was Families 1 and 4 presented to their local dermatol- present on the database of Single Nucleotide Polymor- ogy services, while Families 2 and 3 were identiﬁed phisms (dbSNP), 1000 Genome Project, NHLBI Exome through the International Pachyonychia Congenita Variant Server, Exome Aggregation Consortium Research Registry. All reported PPK since early child- (ExAC) or the Genome Aggregation Database (gno- hood with a positive family history for Patients 2, 3 mAD). and 4. None of the affected individuals had a history

ª 2018 British Association of Dermatologists Clinical and Experimental Dermatology (2019) 44, pp528–534 529 Novel and recurrent mutations in keratin 1 F. J. D. Smith et al.

Table 1 Molecular and clinical details of cases in this study (novel and recurrent) compared with previously reported cases.

Mutation

Protein DNA Keratin Family change change Exon domain Clinical description First reported

1 p.Leu187Pro c.560T>C 1 1A EI. Spontaneous case, with a history of blistering, scaling Current study and erythema since birth. As an adult, diffuse (novel mutation) hyperkeratosis on trunk, limbs and acral sites p.Leu187Phe c.559C>T 1 1A EI. Spontaneous case with erythroderma, erosions, and Szeverenyi et al., 20085 blisters on the entire body surface at birth and (www.interfil.org) palmoplantar and flexural areas of hyperkeratosis in the later stage p.Leu187Phe c.559C>T 1 1A EI. Spontaneous case, generalized hyperkeratosis involving palms and soles 2 p.Leu485Phe c.1453C>T 7 2B PPK. Familial case of diffuse PPK with sharp demarcation Current study developed in early childhood (novel mutation) p.Leu485Pro c.1454T>C 7 2B Generalized severe EI including PPK (mother had EI naevus) Arin et al., 20116 3 p.Ser233Leu c.698C>T 2 1B PPK. Familial case with diffuse well-demarcated plantar Current study (recurrent hyperkeratosis with slightly erythematous borders since mutation) infancy. Fissuring hyperkeratosis of the palms and fingertips p.Ser233Leu c.698C>T 2 1B EPPK (Vorner€ type). Familial case with diffuse PPK with a Szeverenyi et al., 20085 well-circumscribed erythematous margin. No (www.interfil.org) hyperkeratosis affecting other body sites. p.Ser233Leu c.698C>T 2 1B EPPK (Vorner€ type). Familial case with diffuse PPK with a well- circumscribed erythematous margin. No hyperkeratosis affecting other body sites p.Ser233Leu c.698C>T 2 1B EPPK (Vorner€ type). Familial case with diffuse yellowish hyperkeratosis on palms and soles, which developed shortly after birth, with a clear border and demarcated by an erythematous rim p.Ser233Leu c.698C>T 2 1B EPPK (Vorner€ type). Familial case with diffuse yellowish hyperkeratosis on palms and soles, which developed shortly after birth, with a clear border and demarcated by an erythematous rim p.Ser233Leu c.698C>T 2 1B EPPK (Vorner€ type). Developed shortly after birth. Holtz et al., 20168 4 p.Gln418_ c.1254 + 1G>A 6 2B PPK. Familial case with diffuse palmoplantar hyperkeratosis Current study Ile419ins18 that developed shortly after birth (recurrent mutation) p.Gln418_ c.1254 + 1G>A 6 2B EPPK. Familial case, with hyperkeratosis restricted to the Hatsell et al., 20013 Ile419ins18 palms and soles, which developed in childhood. Varying severity between family members from patchy plantar keratoderma to confluent plantar keratoderma. A violaceous erythematous border was observed at the edge of the keratoderma in severe cases p.Gln418_ c.1254 + 1G>A 6 2B EPPK. Familial case, with hyperkeratosis restricted to the Ile419ins18 palms and soles, which developed in childhood. Varying severity between family members from patchy plantar keratoderma to confluent plantar keratoderma. A violaceous erythematous border was observed at the edge of the keratoderma in severe cases p.Gln418_ c.1254 + 1G>A 6 2B EPPK. Familial case, with hyperkeratosis restricted to the Ile419ins18 palms and soles, which developed in childhood. Varying severity between family members from patchy plantar keratoderma to confluent plantar keratoderma. A violaceous erythematous border was observed at the edge of the keratoderma in severe cases

EI, epidermolytic ichthyosis; EPPK, epidermolytic palmoplantar keratoderma; PPK, palmoplantar keratoderma.

530 Clinical and Experimental Dermatology (2019) 44, pp528–534 ª 2018 British Association of Dermatologists Novel and recurrent mutations in keratin 1 F. J. D. Smith et al.

Family 2

(a) (b)

(c)

(d) Wild type K1, exon 7 (e) K1 p.Leu485Phe

Figure 2 Family 2: the proband had diffuse keratoderma of (a,b) plantar and (c) palmar regions. (d,e) Mutation analysis showing DNA sequence (nucleotides 1447–1461) of exon 7 of KRT1 in (d) an unaffected control and (e) the proband. The arrow indicates a heterozygous mutation c.1453C>T leading to the missense mutation p.Leu485Phe.

of collodion membrane, or of hair, teeth or cardiac analysis of this patient identified a previously un- abnormalities. reported heterozygous missense mutation in exon 1 of In Family 1, the proband was a 23-year-old man the keratin 1, p.Leu187Pro; c.560T>C (Fig. 1d,e and who was wheelchair-bound. He presented as an adult Fig. 4). DNA was not available from any other family with diffuse hyperkeratosis on his trunk and limbs in members. This mutation is within the 1A domain of addition to acral sites, having not sought medical keratin 1, and another pathogenic variant has previ- attention for many years. Skin fragility and blistering ously been reported at the same codon, p.Leu187Phe; were reported at sites of trauma, with a history of blis- c.559C>T in two patients with typical EI (Table 1, tering, scaling and erythroderma being present from www.interfil.org).5 birth. Severe flexion contractures of the fingers and The proband of Family 2 was a 39-year-old woman, toes were evident, with surgical correction considered who presented with a history since early childhood of limited in terms of benefit (Fig. 1a–c). Osteopenia, diffuse keratoderma of bilateral palms and plantar feet chronic pain, and the psychological impact of his with sharp demarcation and yellow hue, with milder debilitating skin disease, resulting in social isolation callosities and fissuring affecting the palms and finger- and childhood bullying, necessitated a multidisci- tips (Fig. 2a–c). Her older brother, mother and plinary management approach. There was no known maternal grandmother all had similar findings. A pre- family history, although the proband was estranged viously unreported mutation, p.Leu485Phe; from most of his close relatives so further clinical and c.1453C>T (Fig. 2d,e and Fig. 4) was identified in genetic information was unobtainable. Mutation exon 7 of KRT1 in this patient. The mutation was

ª 2018 British Association of Dermatologists Clinical and Experimental Dermatology (2019) 44, pp528–534 531 Novel and recurrent mutations in keratin 1 F. J. D. Smith et al.

(a) Family 3 K1 p.Ser233Leu (b)

Figure 3 (a,b) Family 3: the proband had diffuse well-demarcated plantar hyperker- (e) (f) atosis with a slightly erythematous border, and was found to have the mutation K1 p.Ser233Leu. Family 4: (c) the proband and her daughter had diffuse hyperkeratosis of the palms, and both had the mutation K1 p.Gln418_Ile419ins18. (d) Peeling of skin on the proband’s palm associated with oral acitretin therapy and (e,f) diffuse hyperkeratosis of the soles with some sparing of the arch.

p.Arg179Pro p.Lys177Asn (2) p.Val176_Lys197del (3) p.Leu486Arg c.591+1G>A; p? p.Leu486Pro (2) p.Leu214Pro c.591+1G>C; p? p.Asn188Thr p.Leu485Pro p.Leu208Pro p.Leu485Phe p.Asp170_Ile173del p.Asn188Lys (2) p.Phe200Leu p.Ile479Phe (2) p.Asn188Ser (6) p.Glu168_Pro171del p.Ile479Thr (9) p.Glu490Gly p.Leu187Phe (2) p.Glu478Gln p.Glu490Gln p.Leu187Pro p.Glu478Lys p.Asp340Val p.Glu490Lys (2) p.Leu161Pro p.Ser186Pro p.Asp340Gly (3) p.Glu478Asp (3) p.Ser233Leu (6) p.Leu437Pro p.Glu489Lys p.Val155Asp p.Met339Lys p.Phe231Leu p.Val623Cysfs*31 (2) p.Val155Gly L1 L12 L2 S p.Gly621Valfs*32 p.Lys74Ile 1A 1B 2A 2B p.Gly585Trpfs*69 p.Gly537Ilefs*77 p.Tyr587Leufs*67 p.Thr481Pro p.Gly543Valfs*70 p.Gln418_Ile419ins (4) p.Tyr482Cys p.Gly519Glufs*94 p.Phe191Ile p.Ala459_Gln466del p.Phe191Leu p.Phe191Cys p.Ser193Pro

Figure 4 Schematic diagram showing the protein domain structure of keratin 1 and previously reported mutations. Those from this study are included; novel mutations are shown in red and recurrent ones in blue.

present in two other affected family members: the demarcated plantar hyperkeratosis with slightly ery- patient’s brother and mother. Another pathogenic mis- thematous borders (Fig. 3a,b). Fissuring hyperkeratosis sense mutation, p.Leu485Pro; c.1454T>C, has previ- of the palms and ﬁngertips was present, but no nail ously been reported at this position in the 2B domain dystrophy, transgrediens or blistering. Her father and of keratin 1 in a patient with PPK (Table 1).6 her paternal grandmother, great-grandmother and In Family 3, the proband was a 43-year-old woman, great-aunt all had similar symptoms. She was found who had a history since infancy of diffuse well- to have a mutation in the 1B domain of KRT1,

532 Clinical and Experimental Dermatology (2019) 44, pp528–534 ª 2018 British Association of Dermatologists Novel and recurrent mutations in keratin 1 F. J. D. Smith et al. p.Ser233Leu; c.698C>T, which has been previously were also supported by a grant from the Pachyonychia reported (www.interfil.org) in several patients with Congenita Project (www.pachyonychia.org) to FJDS. EPPK (Table 1, Fig. 4).5 The proband of Family 4 was a 32-year-old woman, who had diffuse plantar and palmar hyperkeratosis that Learning points peeled significantly once acitretin 20 mg once daily was introduced (Fig. 3c–f). Her grandmother, mother, son • Mutations in keratin genes underlie a variety of and daughter were similarly affected. Genetic analyis epidermal and nonepidermal cell-fragility disor- identified the recurrent pathogenic mutation, ders, and are the genetic basis of a number of the c.1254 + 1G>A, causing a splice site mutation in exon inherited palmoplantar keratodermas. 6, leading to utilization of a novel in-frame splice site 54 • EPPK (OMIM 144200) is an autosomal domin- bases downstream, with subsequent insertion of 18 ant disorder usually caused by mutations in the amino acids into the 2B rod domain, producing the keratin 9 gene (KRT9) but mutations in the ker- mutation, p.Gln418_Ile419ins18, previously reported atin 1 gene (KRT1) have been reported in associ- in association with EPPK (Table 1, Fig. 4).3 ation with this phenotype. Keratin intermediate filaments are characterized by • EI (OMIM 113800), the major keratinopathic tissue-specific expression patterns, with the cell-type ichthyosis, is characterized by congenital erythro- and tissue-specific expression profile of the mutated derma, blistering and erosions of the skin. keratin dictating the cell type and body site that will • Causative mutations in KRT1 and KRT10 have be affected by the disorder.2 KRT1 and KRT10 confer been described in epidermolytic ichthyosis, with structural integrity to suprabasal keratinocytes, and PPK being present primarily in association with mutations in these genes impair the assembly of the the former. keratin cytoskeleton. In some keratin disorders such as • Mutations in KRT1 are not only scattered pachyonychia congenita, keratin mutations are pre- throughout the keratin 1 protein but can result dominantly clustered to the helix boundary domains in a range of phenotypes as further confirmed by at either end of the rod domain. However, the 84 these mutations, giving a complex genotype/ cases previously reported with mutations in KRT1 phenotype pattern. (www.interfil.org) show that mutations in KRT1 are • This report adds to the mutational and clinical not only scattered throughout keratin 1 as shown in spectrum, demonstrating wide phenotypic varia- Fig. 4, but can result in a range of phenotypes6 as fur- tion in KRT1 mutations, despite their unifying ther confirmed by these mutations, giving a complex molecular basis. genotype/phenotype pattern. A wide range in phenotypic severity is also reported in association with KRT1 mutations, particularly in EI.7,8 Mutations in the keratin 9 gene (KRT9) also cause epidermolytic PPK; EPPK. References With the development of new gene and mutation-speci- 1 Moll R, Franke WW, Schiller DL et al. The catalog of fic therapies for inherited disorders of keratinization, it is human cytokeratins: patterns of expression in normal becoming increasingly important to accurately docu- epithelia, tumours and cultures cells. Cell 1982; 31:11– ment both genotype and phenotype. Our cases add to 24. the mutational and clinical spectrum and demonstrate 2 Corden LD, McLean WHI. Human keratin diseases: wide phenotypic variation in KRT1 mutations, despite hereditary fragility of specific epithelial tissues. Exp their unifying molecular basis. Dermatol 1996; 5: 297–307. 3 Hatsell SJ, Eady RA, Wennerstrand L et al. Novel splice site mutation in keratin 1 underlies mild epidermolytic Acknowledgements palmoplantar keratoderma in three kindreds. J Invest Dermatol 2001; 116: 606–9. We thank the patients and their family members for 4 Oji V, Tadini G, Akiyama M et al. Revised nomenclature their cooperation in this study. FJDS, NJW and MZ and classification of inherited ichthyoses: results of the were supported by The Centre for Dermatology and First Ichthyosis Consensus Conference in Soreze 2009. Genetic Medicine at the University of Dundee, which is J Am Acad Dermatol 2010; 63: 607–41. funded by a Wellcome Trust Strategic Award 5 Szeverenyi I, Cassidy AJ, Chung CW et al. The Human (098439/Z/12/Z to WHIMcL), and FJDS and NJW Intermediate Filament Database: comprehensive

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information on a gene family involved in many human associated with mild epidermolytic ichthyosis. Br J diseases. Hum Mutat 2008; 29: 351–60. Dermatol 2010; 162: 875–9. 6 Arin MJ, Oji V, Emmert S et al. Expanding the keratin 8 Holtz A, Oji V, Bourrat E et al. Expanding the clinical and mutation database: novel and recurrent mutations and genetic spectrum of KRT1, KRT2 and KRT10 mutations in genotype-phenotype correlations in 28 patients with keratinopathic ichthyosis. Acta Derm Venereol 2016; 96: epidermolytic ichthyosis. Br J Dermatol 2011; 164: 442–7. 473–8. 7 Bolling MC, Bladergroen RS, van Steensel MAM et al. A novel mutation in the L12 domain of keratin 1 is

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