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Folding the proper action: how insulin get “chaperoned” to find its way out

Insulin is produced in the beta cells of the pancreas. The hormone is produced as a precursor called proinsulin. Insulin needs help acquiring the right structure. For proinsulin to mature into functional insulin, it needs to be folded and processed correctly to acquire the right structure with assistance from proteins that are termed chaperones. The researchers have now discovered and identified such a chaperone, a pro-insulin chaperone termed glucose-regulated protein GRP94. A protein that assists in the process of insulin folding has just been discovered in a new study conducted by researchers at the Department of Biomedical Sciences at University of Copenhagen. They hope the new research results can be used to develop treatments for conditions such as increased level of insulin in the blood (hyperinsulinemia). Even though researchers have been familiar with and studied the hormone insulin for more than a 100 years now, especially related with diabetes, they still make new discoveries concerning this hormone.  

Indeed, researchers from the Faculty of Health and Medical Sciences at the University of Copenhagen have uncovered a hitherto unknown process in the production of insulin. Even though pro-insulin has a relatively short sequence, it still needs help acquiring the right structure (folding) to become mature, functional insulin. However, several other studies have shown that proinsulin can be folded without help from proteins in artificial cell-free conditions. The research conducted in live cells shows, that pro-insulin is not folded correctly and does not acquire the right structure without help from GRP94. Pre-proinsulin is cleaved by signal peptidase to form proinsulin that folds on the luminal side of the endoplasmic reticulum (ER), forming three stable disulfide bonds. Properly folded pro-insulin forms dimers and exits from the endoplasmic reticulum, trafficking through Golgi complex into immature secretory granules. There, C-peptide is proteolytically removed, allowing fully bioactive insulin to ultimately be stored in mature granules for secretion.

In the study the researchers removed or inhibited the protein GRP94 in to see what happened with the pro-insulin and the cells. GRP94 is essential for growth and development of multicellular organisms and is highly expressed in pancreas and bronchial epithelium due to their intense secretory function.  GRP94 expression is upregulated in response to low glucose concentrations and other metabolic stresses, e.g., hypoxia.  Recently, GRP94 has been ablated in pancreatic and duodenal homeobox 1 (Pdx1)-expressing cells and shown to be an essential regulator of β-cell development, mass and function. In other words it is essential for pancreas to produce insulin. Scientists observed that the pro-insulin was not folded correctly and the beta-cells did not secrete sufficient amounts of insulin. Besides, removal of GRP94 did not affect cell viability; in other words, nothing happened to the cells after they had removed the protein. And this was confusing.

GRP94 is indeed instrumental in the initiation of both the innate and adaptive immune response, participates in protein folding, interacts with other components of the ER protein folding machinery, stores calcium and assists in the targeting of misfolded proteins for degradation. Client proteins of GRP94 are selective with critical roles in immunity, growth signaling and cell adhesion. So why its loss would not be so deleterious for beta cells? Scientists do still not know the answer. But the pathway is not so simple: there are entire cellular signalings involved in the process.  For example, one of these is the IRE1-XBP1, which control the folding of pro-insulin under situations of oxidative stress. Furthermore, other molecular partners such as Hsp90, Hsp60, BiP and the protein disulfide isomerase (PDI) are needed. All of these collaborate at the endoplasmic reticulum to assemble proteins, put them in their final three-dimensional configuration and route them to cell compartments or out of the cell.

In case the protein concerned has sulfur bonds, the PDI is actively involved and the XBP-1 regulator is directly responsible for the upstream process because it is a genetic regulator of PDI itself. Actually, other PDI isoforms PDIR, ERp5, ERp44 and ERp46) are involved, because insulin folding and secretion becomes lost once one of these protein is deleted. However, the researchers do not understand why the PERK sensor upstream of the IRE1-PERK-ATF6 pathway does not impair insulin secretion while failing in function. This is in spite of recently identified mechanisms that may protect insulin from oxidation and damage during folding. One of these involve calreticulin, a protein that keeps ionic calcium levels inside the cell in check. The other involves reticulon 3 (RTN-3), a molecular chaperone that directs the elimination of damaged pre-proteins during the folding process. They speculate that a full elucidation of the molecular machinery is going to be needed. Once misteries beneath are solved, informations might be used to develop a “molecular medicine” to restore insulin producion to treat the global burden of diabetes.

  • Edited by Dr. Gianfrancesco Cormaci, PhD, specialist in Clinical Biochemistry.

Scientific references

Argon Y, Bresson SE et al. Cells 2020; 9(8):1844.

Ikezaki M et al. Biochimie. 2020 May; 171-172:1-11.

Mojtaba GS et al. Diabetes 2019 Apr; 68(4):747-60

Tsuchiya Y et al.  Cell Biol 2018; 217(4):1287-1301.

Sowers CR et al. J Biol Chem. 2018; 293(14):5134.

Dott. Gianfrancesco Cormaci
- Laurea in Medicina e Chirurgia nel 1998 (MD Degree in 1998) - Specialista in Biochimica Clinica nel 2002 (Clinical Biochemistry residency in 2002) - Dottorato in Neurobiologia nel 2006 (Neurobiology PhD in 2006) - Ha soggiornato negli Stati Uniti, Baltimora (MD) come ricercatore alle dipendenze del National Institute on Drug Abuse (NIDA/NIH) e poi alla Johns Hopkins University, dal 2004 al 2008. - Dal 2009 si occupa di Medicina personalizzata. - Guardia medica presso strutture private dal 2010 - Detentore di un brevetto sulla preparazione di prodotti gluten-free a partire da regolare farina di frumento immunologicamente neutralizzata (owner of a patent concerning the production of bakery gluten-free products, starting from regular wheat flour). - Responsabile del reparto Ricerca e Sviluppo per la società CoFood s.r.l. (leader of the R&D for the partnership CoFood s.r.l.) - Autore di un libro riguardante la salute e l'alimentazione, con approfondimenti su come questa condizioni tutti i sistemi corporei. - Autore di articoli su informazione medica, salute e benessere sui siti web salutesicilia.com e medicomunicare.it