Over Representation Analysis (ORA)


Case study: ICHTHYOSIS, CYCLIC, WITH EPIDERMOLYTIC HYPERKERATOSIS OMIM #607602
Cyclic ichthyosis, with epidermolytic hyperkeratosis (CIEHK; OMIM #607602) is a subtype of bullous congenital ichthyosiform erythroderma (BCIE).
Based on its clinic-pathological feature – annular and polycyclic erythematous plaques over the proximal extremities and trunk – the disease has been named also as “annular epidermolytic ichthyosis” (Sheth et al., 2007).
CIEHK involves cornification caused by mutations in the keratin 1 gene (KRT1; HGNC:6412; ENSG00000167768) or keratin 10 (KRT10; HGNC:6413, ENSG00000186395) gene.
We performed standard and network-based ORA on the two genes and we retained enriched terms with a p-value < 0.01, after correction with the Benjamini-Hochberg (False Discovery Rate, FDR) procedure. For the sake of clarity, by considering the hierarchical structure of the annotation sources, we report in Tables 1, 2 and 3 only over-represented leaf terms.

Over-represented Biological Processes (GO-BP)
Over the GO-BP resource (Table 1), the standard method highlights processes strictly related to the disease: “peptide cross-linking”, “skin epidermis development”, “cornification” and “keratinization”.
The network-based method adds three new terms: “skin development”, “regulation of keratinocyte proliferation” and “neuromuscular process controlling posture”.
Interestingly, “skin development” is strictly related to the “cornification” process, and “keratinocyte proliferation” has been observed in mouse model for BCIE (Porter et al., 1998). Moreover, looking for the involvement of neuromuscular processes, a psycho-motor retardation (motor skills and language) has been observed in a subject suffering of ichthyosis (because of a mutation in the KRT10 gene; Diociaiuti et al., 2014).

Table 1. Over-represented Biological Processes (GO-BP)

EnrichmentTermN1N2BackgroundFDRDescription
SGO:0098773210156992.37E-05skin epidermis development
SGO:0018149259156994.51E-04peptide cross-linking
SGO:00702682113156991.11E-03cornification
SGO:00314242199156992.08E-03keratinization
N**GO:0050884246172371.01E-03neuromuscular process controlling posture
N**GO:00108372109172373.83E-03regulation of keratinocyte proliferation
N**GO:00435882173172375.81E-03skin development
Enrichment: Standard (S) and Network-based (N) procedure. N** indicates a new enriched term not directly associated to the input gene/proteins;
Term: functional annotation identifier;
N1: input genes/proteins belonging to the term;
N2: genes/proteins associated to the functional term;
Background: number of genes used as background of the Fisher’s exact test;
FDR: p-value corrected by using the Benjamini-Hochberg (False Discovery Rate, FDR) procedure;
Description: brief explanation of the term.


Over-represented Cellular components (GO-CC)
Over the GO-CC resource (Table 2), the standard method enriches the term “cornified envelope”.
The network-based method adds “keratin filament”, and the new term “desmosomes”. Their involvement seems coherent with the features of the pathology, since desmosomes participate in the formation of the “cornified envelope” (Candi et al., 2005).

Table 2. Over-represented Cellular components (GO-CC).

EnrichmentTermN1N2BackgroundFDRDescription
SGO:0001533263120433.23E-04cornified envelope
N**GO:0030057256154766.17E-04desmosome
NGO:00450952276154765.07E-03keratin filament
Enrichment: Standard (S) and Network-based (N) procedure. N** indicates a new enriched term not directly associated to the input gene/proteins;
Term: functional annotation identifier;
N1: input genes/proteins belonging to the term;
N2: genes/proteins associated to the functional term;
Background: number of genes used as background of the Fisher’s exact test;
FDR: p-value corrected by using the Benjamini-Hochberg (False Discovery Rate, FDR) procedure;
Description: brief explanation of the term.


Over-represented Reactome pathways
Over the Reactome database, the standard method highlights the pathway Formation of the cornified envelope (Table 3). As previously reported, its over-representation seems coherent with the features of the disease. Moreover, the network-based enrichment analysis (Table 3) adds the new term “Post-translational modification: synthesis of GPI-anchored proteins”. Interestingly, epidermal-specific impairments of GPI anchor in mice defective of Pig-a gene (essential for the formation of the GPI anchor) include harlequin ichthyosis-like features (Hara-Chikuma et al., 2004).

Table 3. Over-represented Reactome pathways

EnrichmentTermN1N2BackgroundFDRDescription
SR-HSA-68093712129102487.86E-04Formation of the cornified envelope
N**R-HSA-1631252374133995.44E-03Post-translational modification: synthesis of GPI-anchored proteins
Enrichment: Standard (S) and Network-based (N) procedure. N** indicates a new enriched term not directly associated to the input gene/proteins;
Term: functional annotation identifier;
N1: input genes/proteins belonging to the term;
N2: genes/proteins associated to the functional term;
Background: number of genes used as background of the Fisher’s exact test;
FDR: p-value corrected by using the Benjamini-Hochberg (False Discovery Rate, FDR) procedure;
Description: brief explanation of the term.




References
  • Sheth N, Greenblatt D, McGrath JA. (2007) New KRT10 gene mutation underlying the annular variant of bullous congenital ichthyosiform erythroderma with clinical worsening during pregnancy. Br J Dermatol. 157(3):602-4.
  • Porter RM, Reichelt J, Lunny DP, Magin TM, Lane EB. (1998) The relationship between hyperproliferation and epidermal thickening in a mouse model for BCIE. J Invest Dermatol. 110(6):951-7
  • Diociaiuti A, Fortugno P, El Hachem M, Angelo C, Proto V, De Luca N, Martinelli D, Boldrini R, Castiglia D, Zambruno G. (2014) Early immunopathological diagnosis of ichthyosis with confetti in two sporadic cases with new mutations in keratin 10. Acta Derm Venereol. 94(5):579-82
  • Hara-Chikuma M, Takeda J, Tarutani M, Uchida Y, Holleran WM, Endo Y, Elias PM, Inoue S. (2004) Epidermal-specific defect of GPI anchor in Pig-a null mice results in Harlequin ichthyosis-like features. J Invest Dermatol.123(3):464-9.
  • Candi E, Schmidt R, Melino G. (2005) The cornified envelope: a model of cell death in the skin. Nat Rev Mol Cell Biol. 6(4):328-40.