Molecular cloning and characterization of cortical rod protein in the giant freshwater prawn Macrobrachium rosenbergii, a species not forming cortical rod structures in the oocytes

Jul 3, 2007Comparative biochemistry and physiology. Part B, Biochemistry & molecular biology

Cloning and study of the cortical rod protein in the giant freshwater prawn, which does not form these structures in egg cells

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Abstract

A 30-kDa protein related to cortical rod protein was identified in the ovary of the giant freshwater prawn.

The protein identified has a 98% similarity in amino acid sequence to cortical rod protein from kuruma prawns.
Presence of mrCRP mRNA was detected in the ovary at a gonadosomatic index (GSI) of 0.2.
Western blot analysis confirmed a CRP-immunoreactive band at 30 kDa in the ovary with a GSI of 1.6.
Immunocytochemistry revealed mrCRP signals in the ovary at a GSI of 0.9, coinciding with vitellin and lipid accumulation.
mrCRP accumulation in mature oocytes was observed, although it may be masked by larger amounts of vitellin and lipids.

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The formation of cortical rod structures is a characteristic of fully mature oocytes in penaeid prawns, but such structures are absent from oocytes of giant freshwater prawn Macrobrachium rosenbergii. In the present study, we first demonstrated the presence of a 30-kDa protein, which was immunologically related to kuruma prawn cortical rod protein (CRP), in the ovary of giant freshwater prawn, and subsequently purified this protein. Furthermore, a cDNA encoding the CRP-like protein was isolated. Based on the high homology (98%) in the amino acid sequence with kuruma prawn CRP, the 30-kDa protein has been identified as a CRP homologue of giant freshwater prawn, designated mrCRP. The RT-PCR analysis revealed that mrCRP mRNA was present in the ovary from a prawn with a gonadosomatic index (GSI) of 0.2. Western blot analysis revealed the presence of a CRP-immunoreactive band of 30kDa in the ovary with GSI of 1.6. By immunocytochemistry, CRP-immunopositive signals were detected in the ovary with GSI of 0.9, that had started to accumulate considerable amounts of vitellins and lipids in the peripheral cytoplasm. With progress of vitellogenesis, mrCRP was apparently accumulated in the mature oocytes, although it was not detectable, presumably because a relatively small amount of mrCRP was masked with large amounts of vitellin and lipids. In giant freshwater prawn without forming cortical rod structures, our findings indicate that the oocytes produce mrCRP, a homologue of CRP found in penaeid prawns.

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Molecular cloning and characterization of cortical rod protein in the giant freshwater prawn Macrobrachium rosenbergii, a species not forming cortical rod structures in the oocytes

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