Spinous keratinocytes form 4-8 layers of cells in the stratum spinosum (spinous layer) of the interfollicular epidermis (reviewed in Fuchs 1990). While spinous layer keratinocytes are considered to be postmitotic (reviewed in Fuchs 1990), they can still be positive for some of the proliferation markers seen in transit amplifying cells, such as MKI67 and histone H3 phosphorylated on serine residue S11. Currently, generic markers of cell proliferation are not annotated in Reactome. The percentage of spinous layer keratinocytes positive for mitotic markers decreases with age and differs between different skin areas (e.g. plantar skin vs. trunk skin) (Nöske et al. 2016). Spinous layer keratinocytes, similar to basal cells of the epidermis, are connected through desmosomes, Ca2+-activated cell cell junctions.
Keratinocytes of the spinosum layer differentiate into keratinocytes of the granulosum layer. Increased Ca2+ concentrations in the upper layer of epidermis induce transcription of granular cell markers, such as LORICRIN (Dlugosz and Yuspa 1993; Hohl et al. 1991) and FLG (filaggrin) (Dlugosz and Yuspa 1993) in a protein kinase C (PKC)-dependent manner. While Ca2+ also stimulates expression of KRT1 and KRT10, activation of PKC leads to rapid inhibition of KRT1 and KRT10 transcription and reduction of KRT1 mRNA stability. Activation of PKC inhibits EGF signaling. PKC-mediated inhibition of KRT1 and KRT10 transcription requires de novo mRNA and protein synthesis. Candidate PKC isoforms that stimulate differentiation of spinous keratinocytes into granular keratinocytes are four PKCs known to be expressed in cultured keratinocytes: PRKCA, PRKCD, PRKCZ, and PRKCH (Dlugosz and Yuspa 1993).
Besides Ca2+, another factor that stimulates differentiation of spinous into granular keratinocytes is hypoxia. Hypoxia leads to HIF1A and EPAS1 (HIF2A)-mediated stimulation of FLG transcription in a Ca2+ independent and cell confluency-independent manner. Hypoxia also stimulates expression of SPINK5, an inhibitor of epidermal serine proteases that regulates conversion of profilaggrin into monomeric filaggrin. Three putative hypoxia response elements (HREs) are found in the promoter region of the human FLG gene, two of which are functional. One of the two functional HREs is evolutionarily conserved. Direct binding of HIF1A/EPAS1 to these HREs has not been demonstrated. Simultaneous HIF1A and EPAS1 deficiency inhibits terminal differentiation of keratinocytes as well as formation of stratum granulosum and cornified envelope but does not affect MKI67 expression in basal layer keratinocytes or KRT10 expression in stratum spinosum (Wong et al. 2015). ARNT (Aryl hydrocarbon receptor nuclear translocator involved in ceramide synthesis) also positively regulates FLG expression without affecting expression of MKI67 in the basal layer or expression of KRT1 and KRT10 in the spinous layer of epidermis (Wong et al. 2015).
Retinoic acid (RA) inhibits transcription of LORICRIN and FLG in granular cells, even in the presence of high Ca2+ (Hohl et al. 1991; Magnaldo et al. 1992) but has little effect on IVL (involucrin) expression (Magnaldo et al. 1992). Critical cell density is also required for accumulation of LORICRIN mRNA (Hohl et al. 1991).
Inhibition of JNK signaling promotes differentiation of keratinocytes and induces the expression of known suprabasal adhesion proteins such as DSC1, DSC3, DSG1, and LY6D (Gazel et al. 2002). LY6D is specifically expressed in keratinocytes from the stratum spinosum at the RNA level, validated by FISH (Cheng et al. 2018) and by
in situ sequencing (Ganier et al. 2024).
Markers of keratinocytes of the spinous layer are summarized in the "Table of markers of spinous keratinocytes in interfollicular epidermis". Some of the markers, in particular AQP3 and LGLAS7, are described in more detail in the section "Keratinocyte stem cell differentiates into transit amplifying cell in the basal layer of interfollicular epidermis".
FABP5, epidermal fatty acid binding protein also known as E-FABP, is a terminal differentiation marker expressed in upper layers of stratum spinosum and in stratum granulosum (Le et al. 1998). FABP5 is a lipid carrier, prevalently expressed in postmitotic keratinocytes of stratum spinosum and barely detectable in transit amplifying cells and basal cells; FABP5 may positively regulate KRT10 expression through NFKB and JNK signaling (Dallaglio et al. 2013).
Together with KRT10, KRT1 aggregates into thin bundles called tonofilaments (reviewed in Fuchs et al. 1990). IL4 and IL13 suppress expression of KRT1 and KRT10 in keratinocytes of stratum spinosum in atopic dermatitis (Totsuka et al. 2017).
KRT24 is expressed in upper layers of stratum spinosum and required for Ca2+ induced terminal differentiation of keratinocytes (Min et al. 2017). KRT24 is also expressed in upper layers of stratum spinosum in bioengineered skin (Klar et al. 2018).
Table of markers of spinous keratinocytes in interfollicular epidermis. Please note that keratinocytes in CellMarker database and PanglaoDB correspond to spinous keratinocytes in Reactome.
| Marker (protein/RNA) | Literature Reference | CellMarker database – RNA/Protein (Hu et al. 2022) | PanglaoDB – RNA (Franzén et al. 2019) |
|---|
| AQP3 (protein) | Sugiyama et al. 2001
Ma et al. 2002
Sougrat et al. 2002 | No | Yes |
| FAPB5 (protein) | Le et al. 1998
Dallaglio et al. 2013 | No | No |
| KRT1 (protein) | reviewed in Fuchs et al. 1990
Totsuka et al. 2017 | No | Yes |
| KRT10 (protein) | Totsuka et al. 2017 | No | Yes |
| KRT24 (protein) | Min et al. 2017
Klar et al. 2018 | No | No |
| LGALS7 (protein) | Magnaldo et al. 1995
Umayahara et al. 2020 | No | Yes |
| LY6D (RNA) | Gazel et al. 2003
Cheng et al. 2018
Ganier et al. 2024 | No | No |
