The Pancornulins: A Group of Small Proline Rich-Related Cornified Envelope Precursors with Bifunctional Capabilities in Isopeptide Bond Formation

Abstract

In this report, the pancornulins are identified as members of the spr (small, proline-rich) multigene family by amino acid sequence and mass spectrometry analyses. One of the pancornulins (14.9 kDa) is identical to the protein predicted by spr-1 clone 128. The other pancornulins (16.9 kDa and 22 kDa) are novel members of the spr family. Immunoelectron microscopy of purified cornified envelopes with a pancornulin-specific antibody established these proteins more definitively as cornified envelope precursors. In addition, two-dimensional electrophoretic analyses of keratinocyte extracts labeled enzymatically with dansylcadaverine (to identify amine acceptors) or dansylPGGQQIV (to identify amine donors) showed that both glutamine and lysine residues within the pancornulins participate in the isopeptide linkage characteristic of cornified envelope formation. These results contrasted with those obtained using involucrin, a prominent cornified envelope protein shown capable of acting only as an amine acceptor in this system. Novel partial cDNAs obtained after reverse transcription and polymerase chain reaction amplification of total messenger RNA with pancornulin-specific primers suggest that the spr multigene family may be even larger than previously described. The bifunctional reactivity of the pancornulins in cross-linking and the large number of family members identified to date suggest that the pancornulins and other spr-1-related proteins may be more important in cornified envelope formation than previously considered, perhaps functioning as "bridge" molecules during the early phases of cornified envelope assembly.

S100A11, S100A10, Annexin I, Desmosomal Proteins, Small Proline-rich Proteins, Plasminogen Activator Inhibitor-2, and Involucrin Are Components of the Cornified Envelope of Cultured Human Epidermal Keratinocytes

“…Soluble Precursors-Consistent with previous reports (13,14,16,18,35,36), our study identifies involucrin and SPR1A and SPR1B as cytoplasmic, hydrophilic proteins that are incorporated into the cornified envelope. Our results, showing that the central portion of the SPR1A and SPR1B proteins is preferentially released by digestion of cornified envelopes, indicate that SPR1A and SPR1B are cross-linked via the amino-and carboxyl-terminal ends.…”

“…Our results, showing that the central portion of the SPR1A and SPR1B proteins is preferentially released by digestion of cornified envelopes, indicate that SPR1A and SPR1B are cross-linked via the amino-and carboxyl-terminal ends. This suggests that these SPRs may function as molecular cross-bridges to connect two proteins (16). A similar role has been proposed for involucrin (11).…”

“…c Multiple names for the same protein are separated by a slash, some sequences did not find a match in the data base, and blank places indicate that sequence could not be assigned. SPR3 appears to be the 22-kDa pancornulin (16).…”

“…Transglutaminase enzymes catalyze the assembly of this structure via formation of ⑀-(␥-glutamyl)lysine bonds between envelope precursors (4 -7). The proteins that have been identified as constituents of the CE include loricrin (8 -10), involucrin (11)(12)(13)(14), the small proline-rich (SPR) family of proteins (15)(16)(17)(18), cystatin A (19,20), elafin (21)(22)(23), filaggrin (24 -26), keratin (27,28), desmosomal components (27), annexin I (29 -31), plasminogen activator inhibitor-2 (PAI-2) (32), and the 195-kDa and 210-kDa proteins (33,34). Cross-links have been identified within loricrin, elafin, filaggrin, keratin, SPR1, SPR2, and desmoplakin (35).…”

Bricks and mortar of the epidermal barrier

“…Three classes of SPRs have been identified: SPR1 (two members), SPR2 (8-11 members) and SPR3 (1 member) (Kartasova and van de Putte, 1988;Kartasova et al , 1988a;Marvin et al, 1992;An et al, 1993;Gibbs et al, 1993;Greco et al, 1995;Hohl et al, 1995;Austin et al, 1996;Kartasova et al, 1996;Steinert et al, 1998a;Song et al, 1999). The various members of the SPR classes display wide variations of expression in different epithelia.…”