Function of translationally controlled tumor protein (TCTP) in Eudrilus eugeniae regeneration

Abstract

TCTP (Translationally Controlled Tumour Protein) is a multifunctional protein that plays a role in the development, immune system, tumour reversion, and maintenance of stem cells. The mRNA of the Tpt1 gene is over-expressed during liver regeneration. But, the function of the protein in regeneration is not known. To study the role of the protein in regeneration, the earthworm Eudrilus eugeniae was chosen. First, the full length cDNA of the Tpt1 gene was sequenced. The size of the cDNA is 504 bp and the protein has 167 amino acids. The highest level of TCTP expression was documented in the worm after three days of regeneration. The protein was found to be expressed specifically in the epithelial layer of the skin. During regeneration, the protein expression was found to be the highest at the tip of blastema. The pharmacological suppression of TCTP using nutlin-3 and TCTP RNAi experiments resulted in the failure of the regeneration process. The suppression of TCTP caused the arrest of proliferation in posterior amputated worms. The severe cell death was documented in the amputated region of nutlin-3 injected worm. The silencing of TCTP has blocked the modification of clitellar segments. The experiments confirm that TCTP has major functions in the upstream signalling of cell proliferation in the early regeneration process in E. eugeniae.

Understanding the Role of the Clitellum in the Regeneration Events of the Earthworm <b><i>Eudrilus eugeniae</i></b>

“…In the regenerating earthworm, ALP signals are diminished in the clitellum, which may be due to the migration of ALP-positive cells towards the amputation site. Reports suggest that morphological changes were observed in the clitellum during the regeneration process of the earthworm E. eugeniae [Subramanian et al, 2017], which directly supports our findings that it takes place in the clitellum. A BrdU labeling retention assay with regenerating E. eugeniae shows that clumps of BrdU-positive signals were observed in the blastema region [Johnson Retnaraj Samuel et al, 2012], which directly implies cell migration occurs during regeneration.…”

Earthworm, a novel in vivo system to validate antimitotic compounds

“…Earthworms can also be used to study the genes involved in the regeneration of tissues. The role of the TCTP protein in the regeneration of tissues has been shown recently with amputated earthworms and cell division-specific antibodies (Subramanian et al, 2017). In our experiments, all amputated earthworm wounds were healed within 24 h. Regeneration of the blastema was visible 3 days after amputation in control worms.…”

“…Earthworms can also be used to study the genes involved in the regeneration of tissues. The role of the TCTP protein in the regeneration of tissues has been shown recently with amputated earthworms and cell division-specific antibodies (Subramanian et al, 2017).…”

“…To identify whether mitotic cell division was inhibited or not, earthworms injected with water, colchicine, or aqueous extract of rhizomes were subjected to immunohistochemistry (as described by Subramanian et al, 2017) with primary monoclonal antibody of anti-PCNA (Cat. No.…”

“…PCNA is a cell cycle marker protein, expressed only in S (synthesis)phase cells and abundant in actively dividing cells (Foley et al, 1991). Phospho serine 10 histone H3 protein is expressed only in M (mitosis)-phase cells; hence, more of it is found in tissues where mitosis is inhibited (Subramanian et al, 2017). Antibodies of these 2 mitosis-specific proteins were used to find the mitotic cells in the blastema and nearby segments of the regenerating earthworms.…”