A gastrolith protein serving a dual role in the formation of an amorphous mineral containing extracellular matrix.
Despite the proclamation of Lowenstam and Weiner that crustaceans are the "champions of mineral mobilization and deposition of the animal kingdom," relatively few proteins from the two main calcification sites in these animals, i.e., the exoskeleton and the transient calcium storage organs, have been identified, sequenced, and their roles elucidated. Here, a 65-kDa protein (GAP 65) from the gastrolith of the crayfish, Cherax quadricarinatus, is fully characterized and its function in the mineralization of amorphous calcium carbonate (ACC) of the extracellular matrix is demonstrated. GAP 65 is a negatively charged glycoprotein that possesses three predicted domains: a chitin-binding domain 2, a low-density lipoprotein receptor class A domain, and a polysaccharide deacetylase domain. Expression of GAP 65 was localized to columnar epithelial cells of the gastrolith disk during premolt. In vivo administration of GAP 65 dsRNA resulted in a significant reduction of GAP 65 transcript levels in the gastrolith disk. Such gene silencing also caused dramatic structural and morphological deformities in the chitinous-ACC extracellular matrix structure. ACC deposited in these gastroliths appeared to be sparsely packed with large elongated cavities compared with the normal gastrolith, where ACC is densely compacted. ACC spherules deposited in these gastroliths are significantly larger than normal. GAP 65, moreover, inhibited calcium carbonate crystallization in vitro and stabilized synthetic ACC. Thus, GAP 65 is the first protein shown to have dual function, involved both in extracellular matrix formation and in mineral deposition during biomineralization.
Shechter
Assaf
A
Department of Life Sciences, Ben Gurion University of the Negev, Beer-Sheva 84105, Israel.
Glazer
Lilah
L
Cheled
Shira
S
Mor
Eyal
E
Weil
Simy
S
Berman
Amir
A
Bentov
Shmuel
S
Aflalo
Eliahu D
ED
Khalaila
Isam
I
Sagi
Amir
A
D000595:Amino Acid Sequence; D000818:Animals; D003400:Astacoidea; D002119:Calcium Carbonate / Q000737:chemistry*; D002686:Chitin / Q000737:chemistry; D004247:DNA / Q000235:genetics / Q000502:physiology*; D005109:Extracellular Matrix / Q000378:metabolism*; D016326:Extracellular Matrix Proteins / Q000737:chemistry; D020869:Gene Expression Profiling; D020868:Gene Silencing; D008855:Microscopy, Electron, Scanning; D008903:Minerals / Q000737:chemistry*; D008954:Models, Biological; D008969:Molecular Sequence Data; D034622:RNA Interference; D012333:RNA, Messenger / Q000378:metabolism
D016326:Extracellular Matrix Proteins; D008903:Minerals; D012333:RNA, Messenger; D002686:Chitin; D004247:DNA; D002119:Calcium Carbonate