Cell Biology ~ Proteomics & Calcium Biology
1.Our Biology
2.Our Research Technologies
3.Independent Publications
1. Our Biology
Cell Signalling
The mechanisms used to relay messages within cells are of intense biomedical interest. Complex signalling pathways underlie normal development and health of cells. Many diseases are associated with cell signalling anomalies. Numerous drugs target the principal signalling effectors, calcium and protein phosphorylation.
.....................................................
Calcium biology
Calcium has many roles inside and outside cells necessitating strict regulation at different concentrations in various locations. Calcium signals are transmitted through cells as transient increases of calcium which normally is at a very low concentration in the cytosol (cellular fluid). Toxicity arises if these calcium elevations are too large or frequent. It is clear that excess calcium can lead to cell death but disease-related disruptions of calcium signalling are not well understood.
.....................................................
Dental enamel cells
We initiated investigations of calcium handling in enamel cells questioning how they make such a highly-calcified product (tooth enamel is 40% calcium) without suffering the toxic effects of excess intracellular calcium.Of general biomedical value, this research model comprises epithelial cells that have an informatively elongate morphology and undergo functionally-distinct phases of development. Dentally, these cells are also central to the understanding of enamel biomineralization and defects; more information ![]()
.....................................................
Current research focus
- intracellular calcium handling and toxicity avoidance during biomineralization of enamel
- function of calbindins (calcium-binding proteins)
- ERp29 (a novel protein in the endoplasmic reticulum)
- enamel cell proteomes in health and disease
.....................................................
2. Our Research Technologies
Molecular and cellular biology
We are using a broad range of experimental approaches from the DNA level (eg cDNA cloning, TaqMan PCR) through protein (eg 2-D gels, recombinant protein engineering), cellular (eg confocal microscopy) and physiological levels (eg transgenic mouse characterisation).
Proteomics and protein biochemistry
Our speciality is microscale protein biochemistry, a challenging area necessitated by the scarcity of enamel cells. Several novel findings have followed characterisation of proteins expressed by enamel cells ('proteome analysis') [review].
An online database (ToothPrint) of dental proteins has been established.

.....................................................
3. Independent Publications
A. Enamel cell biology – how is bulk calcium handled safely?
We developed microscale proteomic approaches and characterised enamel epithelial cells from developing teeth in neonatal rats and mice. This information was used to investigate the mechanistic basis of calcium transport across enamel cells. Our findings contradicted the classical "calcium ferry" dogma and led to development of a new paradigm for transcellular calcium transport that we've named "calcium transcytosis". Increasingly it appears this organelle-based mechanism could be more generally applicable across biology.
- Mangum, J.E., Veith, P.D., Reynolds, E.C. and Hubbard, M.J. (2006) Towards second-generation proteome analysis of murine enamel-forming cells. Eur. J. Oral Sci. 114, 259-265 (PMID: 16674695)
- Turnbull, C.I., Looi, K., Mangum, J.E., Meyer, M., Sayer, R.J. and Hubbard, M.J. (2004) Calbindin-independence of calcium transport in developing teeth contradicts the calcium-ferry dogma. J. Biol. Chem. 279, 55850-55854 (PMID: 15494408)
- Hubbard, M.J. and Kon, J.C. (2002) Proteomic analysis of dental tissues. J. Chromatogr. B, 771, 211-220 (PMID: 12016000)
- Franklin, I.K., Winz, R.A. and Hubbard, M.J. (2001) Endoplasmic reticulum Ca2+-ATPase pump is up-regulated in calcium-transporting dental enamel cells: A non-housekeeping role for SERCA2b. Biochem. J., 358, 217-224 (PMID: 11485570)
- Hubbard, M.J., Faught, M.J., Carlisle, B.H. and Stockwell, P.A. (2001) ToothPrint, a proteomic database for dental tissues. Proteomics 1, 132-135 (PMID: 11680893)
- Hubbard, M.J. (2000) Calcium transport across the dental enamel epithelium. Crit. Rev. Oral Biol. Med., 11, 437-466 (PMID: 11132765)
- Hubbard, M.J. (1998) Proteomic analysis of enamel cells from developing rat teeth. Big returns from a small tissue. Electrophoresis, 19, 1891-1900 (PMID: 9740049)
- Hubbard, M.J. (1998) Enamel cell biology. Towards a comprehensive biochemical understanding. Conn. Tissue Res., 39, 17-32 (PMID: 11063013)
- Hubbard, M.J. (1996) Abundant calcium homeostasis machinery in rat dental enamel cells. Up-regulation of calcium store proteins during enamel hypermineralization implicates the endoplasmic reticulum in calcium transcytosis. Eur. J. Biochem., 239, 611-623 (PMID: 877470)
- Hubbard, M.J. (1995) Calbindin28kDa and calmodulin are hyperabundant in rat dental enamel cells. Identification of the protein phosphatase calcineurin as a principal calmodulin target and of a secretion-related role for calbindin28kDa. Eur. J. Biochem., 230, 68-79 (PMID: 7601126)
B. Calbindins and calretinin – what do they do?
Calbindins and calretinin have been regarded classically as mobile calcium buffers in the cytosol and consequently they're widely investigated as potential medical targets (e.g. in calcium transport and neurodegeneration). Our investigations of these calcium-binding proteins contradicted this view and instead pointed to a role involving interactions with other proteins. These findings, which lead us to contemplate an alternative role in cell signalling, hold fundamental significance for calbindins as medical targets.
- Turnbull, C.I., Looi, K., Mangum, J.E., Meyer, M., Sayer, R.J. and Hubbard, M.J. (2004) Calbindin-independence of calcium transport in developing teeth contradicts the calcium-ferry dogma. J. Biol. Chem. 279, 55850-55854 (PMID: 15494408)
- Sayer, R.J., Turnbull, C.I. and Hubbard, M.J. (2000) Calbindin28kDa is specifically associated with extra-nuclear constituents of the dense particulate fraction. Cell Tiss. Res., 302, 171-180 (PMID: 11131128)
- Hubbard, M.J. and McHugh, N.J. (1995) Calbindin28kDa and calbindin30kDa (calretinin) are substantially localised in the particulate fraction of rat brain. FEBS Lett., 374, 333-337 (PMID: 7589565)
- Hubbard, M.J. (1995) Calbindin28kDa and calmodulin are hyperabundant in rat dental enamel cells. Identification of the protein phosphatase calcineurin as a principal calmodulin target and of a secretion-related role for calbindin28kDa. Eur. J. Biochem., 230, 68-79 (PMID: 7601126)
- Hubbard, M.J. and Carne, A. (1994) Differential feeding-related regulation of ubiquitin and calbindin9kDa in rat duodenum. Biochim. Biophys. Acta, 1200, 191-6 (PMID: 8031840)
- Hubbard, M.J. (1993) Rapid purification and direct microassay of calbindin9kDa utilizing its solubility in perchloric acid. Biochem. J. 293, 223-7 (PMID: 8392333)
C. ERp29 – what does it do?
We discovered ERp29 during proteomic analysis of rat enamel cells, leading to the naming and first description of this ubiquitous resident of the endoplasmic reticulum (ER). The challenge since has been to figure out its functional role, bioinformatics having offered only limited insight. In a series of pioneering studies, we've gathered a variety of clues that collectively point to a novel "housekeeping" role of general importance, probably as a new type of chaperone. Now linked to a broad array of physiological processes and diseases including cancer, ERp29 holds broad potential as a medical target.
- Das, S., Smith, T.D., Sarma, J.D., Ritzenthaler, J.D., Maza, J., Kaplan, B.E., Cunningham, L.A., Suaud, L., Hubbard, M.J., Rubenstein, R.C., and Koval, M (2009) ERp29 restricts Connexin43 oligomerization in the endoplasmic reticulum. Mol. Biol. Cell 20, 2593-64 (PMID: 19321666)
- Shnyder, S.D., Mangum, J.E. and Hubbard, M.J. (2008) Triplex profiling of functionally distinct chaperones (ERp29/PDI/BiP) reveals marked heterogeneity of the endoplasmic reticulum proteome in cancer. J. Proteome Res. 7, 3364-3372 (PMID: 18598068)
- Hermann, V.M., Cutfield, J.F. and Hubbard, M.J. (2005) Biophysical characterization of ERp29: evidence for a key structural role of Cysteine-125. J. Biol. Chem. 280, 13529-13537 (PMID: 15572350)
- Hubbard, M.J., Mangum, J.E. and McHugh, N.J. (2004) Purification and biochemical characterisation of native ERp29 from rat liver. Biochem. J. 383, 589-598 (PMID: 15500441)
- Macleod, J.C., Sayer, R.J., Lucocq, J.M., and Hubbard, M.J. (2004) ERp29, a general endoplasmic reticulum marker, is highly expressed throughout the brain. J. Comp. Neurol. 477, 29-42 (PMID: 15281078)
- Shnyder, S.D. and Hubbard, M.J. (2002) ERp29 is a ubiquitous resident of the endoplasmic reticulum with a distinct role in secretory protein production. J. Histochem. Cytochem., 50, 557-566 (PMID: 11897809)
- Hubbard, M.J. (2002) Functional proteomics. The goalposts are moving. Proteomics, 2, 1069-1078 (PMID: 12362325)
- Hubbard, M.J., McHugh, N.J. and Carne, D.L. (2000) Isolation of ERp29, a novel endoplasmic reticulum protein, from rat enamel cells: Evidence for a unique role in secretory-protein synthesis. Eur. J. Biochem., 267, 1945-1957 (PMID: 10727933)
- Hubbard, M.J. and McHugh, N.J. (2000) Human ERp29: Isolation, primary structural characterisation and two-dimensional gel mapping. Electrophoresis, 21, 3785-379 (PMID: 11271497)
- Demmer, J., Zhou, C.M. and Hubbard, M.J. (1997) Molecular cloning of ERp29, a novel and widely expressed resident of the endoplasmic reticulum. FEBS Lett., 402, 145-150 (PMID: 9037184)
D. Other reports and articles
- Hubbard, M.J. (2006) Editorial – Hierarchical protein identifications and assignments. J. Proteome Res. 5, 733 (PMID: 16625735)
- Mangum, J.E., Farlie, P.G. and Hubbard, M.J. (2005) Proteomic profiling of facial development in chick embryos. Proteomics 5, 2542-2550 (PMID: 15912509)
- Piotte, C.P., Marshall, C.J., Hubbard, M.J., Collet, C. and Grigor, M.R. (1997) Lysozyme and a-lactalbumin from the milk of a marsupial, the common brush-tailed possum (Trichosurus vulpecula). Biochim. Biophys. Acta, 1336, 235-242 (PMID: 9305795)
- Hubbard, M.J. and McHugh, N.J. (1996) Mitochondrial ATP synthase F1-b-subunit is a calcium-binding protein. FEBS Lett., 391, 323-329 (PMID: 8764999)
- Bird, S.D., Walker, R.J. and Hubbard, M.J. (1994) Altered free calcium transients in pig kidney cells (LLC-PK1) cultured with penicillin/streptomycin. In Vitro Cell Dev. Biol. Anim., 30A, 420-4 (PMID: 7952510)
- Hubbard, M.J. and Cohen, P. (1993) On target with a new mechanism for the regulation of protein phosphorylation. Trends Biochem. Sci 18, 172-7 (PMID: 8392229)
.....................................................