Identification of Alternatively Spliced Novel Transcripts of Neuroserpin Gene Involved in Fenib
Serine protease inhibitors play important roles in homeostasis and disease. Neuroserpin belongs to the serpin superfamily of proteins and is mainly expressed in the brain. Studies link neuroserpin with several diseases like epilepsy, dementia, Alzheimer’s and familial encephalopathy with neuroserpin inclusion bodies (FENIB). FENIB is caused by alteredpolymerization of neuroserpin within the endoplasmic reticulum. Neuroserpin follows an interesting mechanism of action where the reactive center loop interacts with the target protease and inserts itself into the center of β-sheet A. Several critical residues are involved in the loop insertion mechanism and stability of neuroserpin. Mutations leading to the alteredbehavior of neuroserpin have been identified. However, alternatively spliced isoforms of neuroserpin are not well studied. Studying the splicing pattern of neuroserpin is important since these proteins are prone to conformational defects and the introduction or deletion of amino acid sequences is expected to alter the loop insertion dynamics. With the help of a combined methodology involving computational tools and molecular biology techniques, alternatively spliced novel isoforms of human neuroserpin gene were identified. Expression of these novel isoforms was confirmed in the human brain using RT-PCR and DNA sequencing. Using FEX and FGENESH, a novel exon was identified that led to the generation of a transcript with an alternate C-terminal domain. In another case, intron retention led to the introduction of a stop codon and hence generation of a truncated variant that lacked the reactive center loop. To study the structural aspects of these isoforms, MD simulation studies were performed followed by a comparison of RMSF and RMSD values. Results obtained in this study will help in deciphering the role of neuroserpin in health and disease.