Seeing Keratinocyte Proteins Through the Looking Glass of Intrinsic Disorder

International Journal of Molecular Sciences Review Seeing Keratinocyte Proteins through the Looking Glass of Intrinsic Disorder Rambon Shamilov 1 , Victoria L. Robinson 2 and Brian J. Aneskievich 3,* 1 Graduate Program in Pharmacology & Toxicology, Department of Pharmaceutical Sciences, University of Connecticut, 69 North Eagleville Road, Storrs, CT 06269, USA; [email protected] 2 Department of Molecular and Cellular Biology, College of Liberal Arts & Sciences, University of Connecticut, 91 North Eagleville Road, Storrs, CT 06269, USA; [email protected] 3 Department of Pharmaceutical Sciences, School of Pharmacy, University of Connecticut, Storrs, CT 06269, USA * Correspondence: [email protected]; Tel.: +1-860-486-3053 Abstract: Epidermal keratinocyte proteins include many with an eccentric amino acid content (com- positional bias), atypical ultrastructural fate (built-in protease sensitivity), or assembly visible at the light microscope level (cytoplasmic granules). However, when considered through the looking glass of intrinsic disorder (ID), these apparent oddities seem quite expected. Keratinocyte proteins with highly repetitive motifs are of low complexity but high adaptation, providing polymers (e.g., profi- laggrin) for proteolysis into bioactive derivatives, or monomers (e.g., loricrin) repeatedly cross-linked to self and other proteins to shield underlying tissue. Keratohyalin granules developing from liquid– liquid phase separation (LLPS) show that unique biomolecular condensates (BMC) and proteinaceous membraneless organelles (PMLO) occur in these highly customized cells. We conducted bioinfor- matic and in silico assessments of representative keratinocyte differentiation-dependent proteins. This was conducted in the context of them having demonstrated potential ID with the prospect of Citation: Shamilov, R.; Robinson, that characteristic driving formation of distinctive keratinocyte structures. Intriguingly, while ID is V.L.; Aneskievich, B.J. Seeing characteristic of many of these proteins, it does not appear to guarantee LLPS, nor is it required for Keratinocyte Proteins through the Looking Glass of Intrinsic Disorder. incorporation into certain keratinocyte protein condensates. Further examination of keratinocyte- Int. J. Mol. Sci. 2021, 22, 7912. specific proteins will provide variations in the theme of PMLO, possibly recognizing new BMC for https://doi.org/10.3390/ijms22157912 advancements in understanding intrinsically disordered proteins as reflected by keratinocyte biology. Academic Editor: Vladimir Keywords: filaggrin; keratinocyte; liquid–liquid phase separation (LLPS); loricrin; proteinaceous N. Uversky membraneless organelle (PMLO); intrinsically disordered protein (IDP) Received: 24 May 2021 Accepted: 20 July 2021 Published: 24 July 2021 1. Introduction 1.1. Protein Intrinsic Disorder and Keratinocyte Biology Publisher’s Note: MDPI stays neutral In silico, in vitro, and in vivo analyses of intrinsically disordered proteins (IDPs) are with regard to jurisdictional claims in expanding the appreciation [1,2] of their diverse and multiple roles as “hubs” or foci of published maps and institutional affil- iations. intracellular signaling, as scaffolds for protein condensates, and in contributing to other distinctive functions because of their presence as a “conformational ensemble” rather than one fixed three-dimensional structure [3]. This unique conformational characteristic is a consequence of amino acid sequences featuring a biased residue composition with increased polar, charged, and structure-breaking residues as compared to compaction- Copyright: © 2021 by the authors. supporting hydrophobic residues. With increased structural flexibility, IDPs feature prop- Licensee MDPI, Basel, Switzerland. erties distinct from fixed three-dimensional structures including increased regulation by This article is an open access article post-translational modification and greater promiscuity for binding partners. It is these distributed under the terms and conditions of the Creative Commons partner proteins, then, that often confer a specific structural conformation to IDPs via Attribution (CC BY) license (https:// their protein–protein interaction [3]. Despite the importance of signaling hubs and protein creativecommons.org/licenses/by/ scaffolds in the cell biology and differentiation of keratinocytes, we found few reports 4.0/). of purposeful IDP investigations in these cells or, more broadly, in epidermal biology. Int. J. Mol. Sci. 2021, 22, 7912. https://doi.org/10.3390/ijms22157912 https://www.mdpi.com/journal/ijms Int. J. Mol. Sci. 2021, 22, x FOR PEER REVIEW 2 of 25 protein–protein interaction [3]. Despite the importance of signaling hubs and protein scaffolds in the cell biology and differentiation of keratinocytes, we found few reports of purposeful IDP investigations in these cells or, more broadly, in epidermal biology. This apparent disconnect, especially in light of what IDP characteristics could mean for interpreting the function of keratinocyte-specific and keratinocyte-expressed proteins, led us to initiate a synergy of IDP and epidermal keratinocyte biology to fill this knowledge gap. Our premise is that specific investigation of intrinsic disorder (ID) in keratinocyte-expressed proteins could facilitate the understanding of their function in ways heretofore not considered. Our approach utilized two complementary avenues: (i) an overview of Int. J. Mol. Sci. 2021, 22, 7912 the keratinocyte literature for IDP-relevant reports, and (ii) an extensive2 of 24 targeted bioin- formatic assessment of ID in keratinocyte proteins, especially for those proteins contributing to keratinocyte-unique structures. We aimed to establish relevance for more exten- Thissive apparent IDP-directed disconnect, studies especially in incutaneous light of what resear IDP characteristicsch and, at could the meansame for time, inform IDP re- interpreting the function of keratinocyte-specific and keratinocyte-expressed proteins, led ussearchers to initiate aregarding synergy of IDPthe andopportunities epidermal keratinocyte for discov biologyery toin fillthe this specialized knowledge tissue of the epider- gap.mis. Our premise is that specific investigation of intrinsic disorder (ID) in keratinocyte- expressed proteins could facilitate the understanding of their function in ways heretofore not considered. Our approach utilized two complementary avenues: (i) an overview of the1.2. keratinocyte Epidermal literature Specialization for IDP-relevant and Keratino reports, andcyte-Related (ii) an extensive Protein targeted Intrinsic bioinfor- Disorder matic assessmentThe epidermis of ID in keratinocyteis the upper-most proteins, especially compartmen for thoset proteinsof the contributingskin. Due to the carefully reg- to keratinocyte-unique structures. We aimed to establish relevance for more extensive IDP-directedulated, progressive studies in cutaneous maturation research of and, its atmajor the same cell time, type, inform the IDP keratinocyte, researchers the epidermis pro- regardingvides an the essential opportunities barrier for discovery function in theto protec specializedt the tissue underlying of the epidermis. dermis and deeper tissues [4]. Within the epidermis (Figure 1), keratinocytes form multiple layers (strata). For the epi- 1.2. Epidermal Specialization and Keratinocyte-Related Protein Intrinsic Disorder dermisThe epidermis in humans is the and upper-most other mammals, compartment ofthese the skin. are Dueclassically to the carefully recognized regu- as the basal layer, lated,mitotically progressive active maturation and of in its majordirect cell contact type, the keratinocyte,with the underlying the epidermis provides dermis; then, the spinous anlayer, essential post-mitotic barrier function keratinocytes to protect the underlying with numerous dermis and deeper cell–cell tissues adhesion [4]. Within points (desmosome the epidermis (Figure1), keratinocytes form multiple layers (strata). For the epidermis in humans“spines”); and other the granular mammals, these layer are characterized classically recognized by keratohyalin as the basal layer, aggregates mitotically involved in keratin activeintermediate and in direct filament contact withreorganization; the underlying and, dermis; ulti then,mately, the spinous the cornified layer, post- layer, exposed to the mitoticenvironment keratinocytes and with composed numerous cell–cellof flattened adhesion kera pointstinocytes (desmosome (squames) “spines”); thefor which nuclear deg- granularradation layer and characterized extensive by protein–protein keratohyalin aggregates cross-lin involvedking in keratin have intermediate taken place in their transition filament reorganization; and, ultimately, the cornified layer, exposed to the environment andfrom composed granular of flattened to cornified keratinocytes (Figure (squames) 1). These for which layers nuclear are degradation recognized and by their histological extensiveposition protein–protein and specific cross-linking proteins derived have taken from place instrata- their transition and keratinocyte-specific from granular gene expres- tosion cornified from (Figurebasal1 to). Thesespinous layers to are granular recognized cells. by theirDue histological to their key position roles and in normal cell physiol- specific proteins derived from strata- and keratinocyte-specific gene expression from basal toogy spinous and tocertain granular disease cells. Due states, to their the key tertiary

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