appeared as a rhombohedron of calcite. Various workers (Okazaki 1960; Beniash
et al. 1997) demonstrated a putative surface “envelope” composed of organic
material, and Benson et al. (1983) gave clear morphological evidence for a network
of occluded organic molecules in the demineralized spicule. Gentle etching of
fractured surfaces revealed a lamellar organization of the mineral, much like
growth rings of a section of a tree trunk (Seto et al. 2004). It was proposed that
these lamellae could be formed from the periodic deposition of additional layers of
mineral that are responsible for increase in girth of the spicule during its development, but there is no direct evidence for that proposal (Seto et al. 2004).
Using affinity-purified antibodies against SM30 and SM50 Seto et al. (2004)
showed by immunoelectron microscopy that fractured, etched surfaces were specifically labeled, thereby providing direct evidence for occlusion of these matrix
proteins. Both these proteins, especially SM50, are also found on the external
surface of the spicule. The picture that emerged is one of well (but not perfectly)
aligned domains of calcite (Berman et al. 1993) in which a fibrous network of
occluded matrix proteins traverse boundaries between domains.
One goal of research on biomineralization is the identification, enumeration, and
function of organic molecules found on, or occluded within, the mineralized
structures. Polysaccharides and proteins dominate the lists in the various
biomineralized tissues that have been closely examined. Application of the methods
of molecular biology has been especially helpful since occluded matrix proteins
often resist routine methods of protein purification and characterization. An
extended discussion of the approaches and results can be found in reviews by
Wilt and Ettensohn (2007) and Killian and Wilt (2008). Suffice it to say that even
though the protein content of the sea urchin spicule is very low (~0.1% by mass,
Wilt 1999), there are apparently over 40 different proteins of which most, but not
all, are acidic and glycosylated
1 . Only one of these 40 has been subjected to a
rigorous test of its functional role: synthesis of SM50 during embryonic development is essential for spicule formation (Peled-Kamar et al. 2002; Wilt et al. 2008a).
While SM50 is necessary for spicule formation, the exact nature of the role SM50
plays has not been elucidated. It might be instructive to engineer and express
counterfeit versions of SM50 to see if particular portions of SM50 can act as a
“dominant negative”.
The genome of S. purpuratus has been sequenced and annotated, and an enumeration of genes known to be involved in spicule formation and/or structure
compiled (Livingston et al. 2006). These proteins are mostly acidic, glycosylated,
secreted and contain a C-lectin domain. The necessity and/or function of any of
them, except SM50, is unknown. We should also remember that proteins important
for spicule formation may not necessarily end up occluded in the composite, e.g.,
the apparent important role of metalloproteases.
1
Recent proteomic work demonstrated over 200 proteins in the spicule (Mann et al. 2010).
7 Molecular Aspects of Biomineralization of the Echinoderm Endoskeleton
207
et al. 1997) demonstrated a putative surface “envelope” composed of organic
material, and Benson et al. (1983) gave clear morphological evidence for a network
of occluded organic molecules in the demineralized spicule. Gentle etching of
fractured surfaces revealed a lamellar organization of the mineral, much like
growth rings of a section of a tree trunk (Seto et al. 2004). It was proposed that
these lamellae could be formed from the periodic deposition of additional layers of
mineral that are responsible for increase in girth of the spicule during its development, but there is no direct evidence for that proposal (Seto et al. 2004).
Using affinity-purified antibodies against SM30 and SM50 Seto et al. (2004)
showed by immunoelectron microscopy that fractured, etched surfaces were specifically labeled, thereby providing direct evidence for occlusion of these matrix
proteins. Both these proteins, especially SM50, are also found on the external
surface of the spicule. The picture that emerged is one of well (but not perfectly)
aligned domains of calcite (Berman et al. 1993) in which a fibrous network of
occluded matrix proteins traverse boundaries between domains.
One goal of research on biomineralization is the identification, enumeration, and
function of organic molecules found on, or occluded within, the mineralized
structures. Polysaccharides and proteins dominate the lists in the various
biomineralized tissues that have been closely examined. Application of the methods
of molecular biology has been especially helpful since occluded matrix proteins
often resist routine methods of protein purification and characterization. An
extended discussion of the approaches and results can be found in reviews by
Wilt and Ettensohn (2007) and Killian and Wilt (2008). Suffice it to say that even
though the protein content of the sea urchin spicule is very low (~0.1% by mass,
Wilt 1999), there are apparently over 40 different proteins of which most, but not
all, are acidic and glycosylated
1 . Only one of these 40 has been subjected to a
rigorous test of its functional role: synthesis of SM50 during embryonic development is essential for spicule formation (Peled-Kamar et al. 2002; Wilt et al. 2008a).
While SM50 is necessary for spicule formation, the exact nature of the role SM50
plays has not been elucidated. It might be instructive to engineer and express
counterfeit versions of SM50 to see if particular portions of SM50 can act as a
“dominant negative”.
The genome of S. purpuratus has been sequenced and annotated, and an enumeration of genes known to be involved in spicule formation and/or structure
compiled (Livingston et al. 2006). These proteins are mostly acidic, glycosylated,
secreted and contain a C-lectin domain. The necessity and/or function of any of
them, except SM50, is unknown. We should also remember that proteins important
for spicule formation may not necessarily end up occluded in the composite, e.g.,
the apparent important role of metalloproteases.
1
Recent proteomic work demonstrated over 200 proteins in the spicule (Mann et al. 2010).
7 Molecular Aspects of Biomineralization of the Echinoderm Endoskeleton
207