Altered Proglucagon Processing in an α-Cell Line Derived from Prohormone Convertase 2 Null Mouse Islets

The endoproteolytic processing of proproteins in the secretory pathway depends on the expression of selected members of a family of subtilisin-like endoproteases known as the prohormone convertases (PCs). The main PC family members expressed in mammalian neuroendocrine cells are PC2 and PC1/3. The differential processing of proglucagon in pancreatic α-cells and intestinal L cells leads to production of distinct hormonal products with opposing physiological effects from the same precursor. Here we describe the establishment and characterization of a novel α-cell line (αTC-ΔPC2) derived from PC2 homozygous null animals. The αTC-ΔPC2 cells are shown to be similar to the well characterized αTC1–6 cell line in both morphology and overall gene expression. However, the absence of PC2 activity in αTC-ΔPC2 leads to a complete block in the production of mature glucagon. Surprisingly, αTC-ΔPC2 cells are able to efficiently cleave the interdomain site in proglucagon (KR 70–71). Further analysis reveals that αTC-ΔPC2 cells, unlike αTC1–6 cells, express low levels of PC1/3 that lead to the generation of glicentin as well as low amounts of oxyntomodulin, GLP-1, truncated GLP-1, and N-terminally extended GLP-2. We conclude that αTC-ΔPC2 cells provide additional evidence for PC2 as the major convertase in α-cells leading to mature glucagon production and provide a robust model for further analysis of the mechanisms of proprotein processing by the prohormone convertases. The endoproteolytic processing of proproteins in the secretory pathway depends on the expression of selected members of a family of subtilisin-like endoproteases known as the prohormone convertases (PCs). The main PC family members expressed in mammalian neuroendocrine cells are PC2 and PC1/3. The differential processing of proglucagon in pancreatic α-cells and intestinal L cells leads to production of distinct hormonal products with opposing physiological effects from the same precursor. Here we describe the establishment and characterization of a novel α-cell line (αTC-ΔPC2) derived from PC2 homozygous null animals. The αTC-ΔPC2 cells are shown to be similar to the well characterized αTC1–6 cell line in both morphology and overall gene expression. However, the absence of PC2 activity in αTC-ΔPC2 leads to a complete block in the production of mature glucagon. Surprisingly, αTC-ΔPC2 cells are able to efficiently cleave the interdomain site in proglucagon (KR 70–71). Further analysis reveals that αTC-ΔPC2 cells, unlike αTC1–6 cells, express low levels of PC1/3 that lead to the generation of glicentin as well as low amounts of oxyntomodulin, GLP-1, truncated GLP-1, and N-terminally extended GLP-2. We conclude that αTC-ΔPC2 cells provide additional evidence for PC2 as the major convertase in α-cells leading to mature glucagon production and provide a robust model for further analysis of the mechanisms of proprotein processing by the prohormone convertases. Since the discovery of propeptide processing in the production of mature insulin (1Steiner D.F. Cunningham D. Spigelman L. Aten B. Science. 1967; 157: 697-700Google Scholar), endoproteolytic cleavage of cellular proteins to generate biologically active products has become recognized as a widespread and fundamental regulatory mechanism of the proteome. This type of processing occurs in multiple subcellular compartments, including the cytoplasm, various organelles, and on membrane surfaces (2Wilkinson K.D. Annu. Rev. Nutr. 1995; 15: 161-189Google Scholar, 3Nakayama K. Biochem. J. 1997; 327: 625-635Google Scholar, 4Martoglio B. Dobberstein B. Trends Cell Biol. 1998; 8: 410-415Google Scholar, 5Sakai J. Rawson R.B. Espenshade P.J. Cheng D. Seegmiller A.C. Goldstein J.L. Brown M.S. Mol. Cell. 1998; 2: 505-514Google Scholar, 6Thornberry N.A. Lazebnik Y. Science. 1998; 281: 1312-1316Google Scholar). Consequent to initial findings that proinsulin processing occurs in the regulated secretory pathway, multiple small peptide hormones and neuropeptides are now recognized as being derived from larger precursors by similar processing in post-Golgi compartments (7Steiner D.F. Kemmler W. Tager H.S. Peterson J.D. Fed. Proc. 1974; 33: 2105-2115Google Scholar, 8Seidah N.G. Chretien M. Day R. Biochimie (Paris). 1994; 76: 197-209Google Scholar, 9Rouille Y. Duguay S.J. Lund K. Furuta M. Gong Q. Lipkind G. Oliva Jr., A.A. Chan S.J. Steiner D.F. Front. Neuroendocrinol. 1995; 16: 322-361Google Scholar, 10Creemers J.W. Jackson R.S. Hutton J.C. Semin. Cell Dev. Biol. 1998; 9: 3-10Google Scholar, 11Seidah N.G. Day R. Marcinkiewicz M. Chretien M. Ann. N. Y. Acad. Sci. 1998; 839: 9-24Google Scholar, 12Viale A. Ortola C. Hervieu G. Furuta M. Barbero P. Steiner D.F. Seidah N.G. Nahon J.L. J. Biol. Chem. 1999; 274: 6536-6545Google Scholar, 13Zhou A. Webb G. Zhu X. Steiner D.F. J. Biol. Chem. 1999; 274: 20745-20748Google Scholar). Early studies of these processing events led to the recognition that peptide cleavage usually occurs C-terminal to dibasic amino acid motifs, followed by removal of the C-terminal basic amino acids by carboxypeptidases, such as carboxypeptidase E (14Fricker L. Annu. Rev. Physiol. 1988; 50: 309-321Google Scholar). Following the discovery of the yeast calcium-dependent subtilisin-like endoprotease kexin (Kex2p), which acts on dibasic motifs in the α mating factor and killer toxin precursors in the distal yeast secretory pathway (15Julius D. Brake A. Blair L. Kunisawa R. Thorner J. Cell. 1984; 37: 1075-1089Google Scholar, 16Fuller R.S. Sterne R.E. Thorner J. Annu. Rev. Physiol. 1988; 50: 345-362Google Scholar, 17Fuller R.S. Brake A. Thorner J. Proc. Natl. Acad. Sci. U. S. A. 1989; 86: 1434-1438Google Scholar), the cloning and characterization of related mammalian enzymes soon followed (18Smeekens S.P. Steiner D.F. J. Biol. Chem. 1990; 265: 2997-3000Google Scholar, 19Smeekens S.P. Avruch A.S. LaMendola J. Chan S.J. Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1991; 88: 340-344Google Scholar, 20Seidah N.G. Marcinkiewicz M. Benjannet S. Gaspar L. Beaubien G. Mattei M.G. Lazure C. Mbikay M. Chretien M. Mol. Endocrinol. 1991; 5: 111-122Google Scholar). These proteases have been designated subtilisin-like prohormone or proprotein convertases (SPCs, or more simply PCs) 1The abbreviations used are: PC, prohormone convertase; GLP-1, -2, glucagon-like peptides 1 and 2; Tricine, N-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]glycine; RT, reverse transcription; MCA, 7-methoxycoumarin; MOPS, 4-morpholinepropanesulfonic acid; SSPE, saline/sodium phosphate/EDTA. 1The abbreviations used are: PC, prohormone convertase; GLP-1, -2, glucagon-like peptides 1 and 2; Tricine, N-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]glycine; RT, reverse transcription; MCA, 7-methoxycoumarin; MOPS, 4-morpholinepropanesulfonic acid; SSPE, saline/sodium phosphate/EDTA. in recognition of their role in processing, not only prohormones, but a wide variety of other precursor proteins that traverse the secretory pathway. Although the mammalian PC family is now quite large, consisting of seven members with varying basic residue specificities, those mainly responsible for precursor processing in the regulated secretory pathway are now believed to be PC3 (also called PC1, and here referred to as PC1/3), PC2 (13Zhou A. Webb G. Zhu X. Steiner D.F. J. Biol. Chem. 1999; 274: 20745-20748Google Scholar), and to a lesser extent PC5/6A (21Barbero P. Rovere C. De Bie I. Seidah N. Beaudet A. Kitabgi P. J. Biol. Chem. 1998; 273: 25339-25346Google Scholar, 22Cain B.M. Vishnuvardhan D. Beinfeld M.C. Peptides. 2001; 22: 1271-1277Google Scholar, 23Dey A. Norrbom C. Xhu X. Stein J. Zhang C. Ueda K. Steiner D.F. Endocrinology. 2004; 145: 1961-1971Google Scholar). The PCs are all calcium-dependent serine endoproteases with acidic pH optima consistent with their function in the calcium-rich acidic secretory granules of neuroendocrine cells. A recent solution of the crystal structure of the PC furin confirms their structural relationship to the subtilase family and provides more detailed insight into the structural basis of substrate recognition (24Henrich S. Cameron A. Bourenkov G.P. Kiefersauer R. Huber R. Lindberg I. Bode W. Than M.E. Nat. Struct. Biol. 2003; 10: 520-526Google Scholar). Like subtilisin, these proteases become active by autocatalytic cleavage of an N-terminal propeptide, which is required for folding of the proenzymes (10Creemers J.W. Jackson R.S. Hutton J.C. Semin. Cell Dev. Biol. 1998; 9: 3-10Google Scholar, 25Hu Z. Zhu X. Jordan F. Inouye M. Biochemistry. 1994; 33: 562-569Google Scholar). A downstream domain of about 150 amino acids, called the P- or Homo B-domain (8Seidah N.G. Chretien M. Day R. Biochimie (Paris). 1994; 76: 197-209Google Scholar, 9Rouille Y. Duguay S.J. Lund K. Furuta M. Gong Q. Lipkind G. Oliva Jr., A.A. Chan S.J. Steiner D.F. Front. Neuroendocrinol. 1995; 16: 322-361Google Scholar), is also required for folding and activity. This domain plays a regulatory role, influencing both the calcium dependence and pH optima (26Zhou A. Martin S. Lipkind G. LaMendola J. Steiner D.F. J. Biol. Chem. 1998; 273: 11107-11114Google Scholar). The variable C-terminal regions of the PCs are less conserved and play a role in their subcellular localization (10Creemers J.W. Jackson R.S. Hutton J.C. Semin. Cell Dev. Biol. 1998; 9: 3-10Google Scholar, 27Steiner D.F. Curr. Opin. Chem. Biol. 1998; 2: 31-39Google Scholar). Although PC1/3 and PC2 share many biochemical and functional characteristics, subtle structural differences must account for observed differences in their recognition of distinct dibasic amino acid motifs within precursors (8Seidah N.G. Chretien M. Day R. Biochimie (Paris). 1994; 76: 197-209Google Scholar, 9Rouille Y. Duguay S.J. Lund K. Furuta M. Gong Q. Lipkind G. Oliva Jr., A.A. Chan S.J. Steiner D.F. Front. Neuroendocrinol. 1995; 16: 322-361Google Scholar, 13Zhou A. Webb G. Zhu X. Steiner D.F. J. Biol. Chem. 1999; 274: 20745-20748Google Scholar). This functional refinement between PC1/3 and PC2 becomes strikingly apparent in the processing of proglucagon, a multifunctional precursor that contains multiple cleavage sites recognized with varying efficiencies by these two convertases (28Rouille Y. Westermark G. Martin S.K. Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 3242-3246Google Scholar, 29Rouille Y. Martin S. Steiner D.F. J. Biol. Chem. 1995; 270: 26488-26496Google Scholar, 30Rouillé Y. Bianchi M. Irminger J.-C. Halban P. FEBS Lett. 1997; 413: 119-123Google Scholar, 31Rouillé Y. Kantengwa S. Irminger J.-C. Halban P.A. J. Biol. Chem. 1997; 272: Scholar). proglucagon be in cells to of products with in the pancreatic α-cell expression of PC2 is with the production of glucagon (28Rouille Y. Westermark G. Martin S.K. Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 3242-3246Google Scholar, 30Rouillé Y. Bianchi M. Irminger J.-C. Halban P. FEBS Lett. 1997; 413: 119-123Google Scholar), in intestinal L cells processing, mainly by leads to the production of glucagon-like peptides 1 and and Y. Martin S. Steiner D.F. J. Biol. Chem. 1995; 270: 26488-26496Google Scholar, 31Rouillé Y. Kantengwa S. Irminger J.-C. Halban P.A. J. Biol. Chem. 1997; 272: Scholar). We here the and characterization of a pancreatic α-cell line from PC2 null M. A. Y. R. M. L. Furuta Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1997; Scholar), which has the to proglucagon processing in a of α-cells in in the absence of PC2 activity. amino acids, a from Lindberg of The a from J. of the of the cell and for from The and from The for from The and membrane from The and to glucagon from The for and the and from the αTC-ΔPC2 Cell PC2 null M. A. Y. R. M. L. Furuta Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1997; to the in the pancreatic α-cells by of A.C. S. S. D. D. 1990; Scholar). The PC2 to PC2 homozygous null to PC2 homozygous animals. by analysis as M. A. Y. R. M. L. Furuta Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1997; Scholar, A.C. S. S. D. D. 1990; Scholar). These of and the as in between and in and used to α-cell A.C. S. S. D. D. 1990; Scholar). with and in and a of the and with into to for to of as of of the cells The cells to well and for more cells in for and The (αTC-ΔPC2) cells here derived from a on and on an or in followed by processing for as M.S. C. Steiner D.F. Scholar). Cell and and cells in with 1 and and as A. Xhu X. R. Stein J. Steiner D.F. J. Biol. Chem. 2003; Scholar). in of and in both cell and cell as well as peptides from cell and in and the in from cell type as well as pancreatic for and and of of 1 of for and with the used of of the by for followed by with 1 and and 1 for and for of the PC2 the used for the type PC2 these to the that to PC2 the products in and and a The and reverse and and proinsulin and and proinsulin 2: and C. PC2 of cell in from cell type varying from to in a of the A. Lindberg I. R.E. The Scholar), in the absence or of the PC2 peptide amino X. Y. Steiner D.F. Lindberg I. Proc. Natl. Acad. Sci. U. S. A. Scholar), a of The for and the of from the substrate as a of cleavage in with 1 acid as X. Y. Steiner D.F. Lindberg I. Proc. Natl. Acad. Sci. U. S. A. Scholar). from cell type amounts of in and and for to in a solution SSPE, solution and and in the same solution in the of in a in for followed by for to more removal of to with a membrane in a solution of with for and with The used as by and for and αTC-ΔPC2 cells for analysis as J. Webb G. Y. Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 2003; Scholar). which in and an a neuroendocrine cell line of active PC2 and to the of PC2 in the processing of proglucagon in the pancreatic an α-cell line PC2 activity The is known to pancreatic a expression of the from the glucagon M. A. Y. R. M. L. Furuta Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1997; Scholar, S. G. M. D. D. 1988; Scholar). to the PC2 null M. A. Y. R. M. L. Furuta Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1997; that PC2 homozygous null and the and A cell line from the pancreatic α-cell that characterized This cell line has been designated αTC-ΔPC2 cells similar to those of the well characterized αTC1–6 cell line A and analysis αTC1–6 cells, αTC-ΔPC2 cells secretory granules with and αTC-ΔPC2 a of to αTC1–6 and further the cell the expression of pancreatic by Like αTC1–6 cells, αTC-ΔPC2 cells express proglucagon their to α-cells Like αTC1–6 and unlike the line αTC-ΔPC2 expression of proinsulin 1 or proinsulin the PC2 null a but active PC2 M. A. Y. R. M. L. Furuta Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1997; and the used in analysis the of all cell a PC2 that is of the same we have not levels of in αTC1–6 or αTC-ΔPC2 cells not the well characterized αTC1–6 α-cell the αTC-ΔPC2 cells express known to be in α-cells and and to express known not to be expressed in α-cells 1 and and that the cell line has α-cell cell line that the PC2 in the αTC-ΔPC2 cells to the null analysis The from the null is the for PC2 in and αTC-ΔPC2 cells as and in all cells express the for the PC2 null a consistent with derived cells and cells, used as not express levels of consistent with from both α-cell express levels of proglucagon, unlike cells. αTC-ΔPC2 of from PC2 homozygous null have that the null leads to the production of a of M. A. Y. R. M. L. Furuta Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1997; Scholar). This is not and must be M. A. Y. R. M. L. Furuta Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1997; Scholar). the production of PC2 on cell from and αTC-ΔPC2 cells. and αTC1–6 cells levels of mature PC2 and levels of as (28Rouille Y. Westermark G. Martin S.K. Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 3242-3246Google The αTC-ΔPC2 cells, a truncated with lesser amounts of or of which to active of levels differences between the PC2 null line and other neuroendocrine cell of a of analysis of from and cell for PC2 activity levels of activity in both αTC1–6 and activity is by the peptide A. Lindberg I. R.E. The Scholar, I. C. Biochemistry. 1995; Scholar, Y. J. Lindberg I. J. 1999; Scholar). the other αTC-ΔPC2 PC2 similar to from cells. These that the αTC-ΔPC2 cell line an α-cell in the absence of active PC2 and provides an model of α-cell function for the of propeptide processing in the absence of PC2 activity. in αTC-ΔPC2 processing of proglucagon is a regulated and leading to the production of of peptide on cell type and of expressed convertases (28Rouille Y. Westermark G. Martin S.K. Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 3242-3246Google Scholar, 29Rouille Y. Martin S. Steiner D.F. J. Biol. Chem. 1995; 270: 26488-26496Google Scholar, 30Rouillé Y. Bianchi M. Irminger J.-C. Halban P. FEBS Lett. 1997; 413: 119-123Google Scholar, 31Rouillé Y. Kantengwa S. Irminger J.-C. Halban P.A. J. Biol. Chem. 1997; 272: Scholar). of cellular by for proglucagon and peptide products that αTC1–6 cells efficiently proglucagon to mature glucagon with amounts of glicentin and αTC-ΔPC2 cells mature but amounts of This from the complete block in proglucagon processing in pancreatic of PC2 null M. A. Webb G. R. M. L. Steiner D.F. J. Biol. Chem. 2001; Scholar), the in αTC-ΔPC2 cells of PC not analysis of peptides that glicentin is a major with amounts of proglucagon in αTC-ΔPC2 cells both and mature glucagon is not cellular the contains a small but of from the αTC-ΔPC2 cells, or processing the dibasic site the processing of the interdomain cleavage site and led to production of glicentin and oxyntomodulin, but not mature from the N-terminal domain of the proglucagon in αTC-ΔPC2 cells. processing of the C-terminal domain of the proglucagon in the αTC-ΔPC2 cells, production to GLP-1, we the of both and truncated and low levels in from αTC-ΔPC2 cells the same cell we but truncated processing the we the generation of N-terminally extended low levels in αTC-ΔPC2 cells αTC1–6 cells by mature glucagon with amounts of glicentin and and small amounts of as well as and extended but truncated and The that cleavage the interdomain and to glicentin and the dibasic and and to generate oxyntomodulin, and the N-terminally extended of and and in the absence of PC2 activity in the αTC-ΔPC2 cells. These processing events are also consistent with the expression of convertase in the αTC-ΔPC2 cells. PC1/3 has been shown to be of these sites Y. Martin S. Steiner D.F. J. Biol. Chem. 1995; 270: 26488-26496Google Scholar, 31Rouillé Y. Kantengwa S. Irminger J.-C. Halban P.A. J. Biol. Chem. 1997; 272: Scholar), we the that PC1/3 is expressed in these cells. analysis of from and cells that αTC-ΔPC2 cells but levels of αTC1–6 cells of PC1/3 expression and cells robust expression of PC1/3 S.P. Avruch A.S. LaMendola J. Chan S.J. Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1991; 88: 340-344Google Scholar, 20Seidah N.G. Marcinkiewicz M. Benjannet S. Gaspar L. Beaubien G. Mattei M.G. Lazure C. Mbikay M. Chretien M. Mol. Endocrinol. 1991; 5: 111-122Google Scholar, A. Martin S. Lipkind G. LaMendola J. Steiner D.F. J. Biol. Chem. 1998; 273: 11107-11114Google Scholar). from and cells PC1/3 expression. the of PC1/3 in the αTC-ΔPC2 cells, for PC1/3 on from the cell (28Rouille Y. Westermark G. Martin S.K. Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 3242-3246Google and the αTC1–6 PC1/3 and cells levels of mature PC1/3. with the low but levels of PC1/3 in from these cells low levels of mature PC1/3 is that PC1/3 also for the low of cleavage of substrate in αTC-ΔPC2 cell and both the observed interdomain as well as the products of proglucagon of αTC1–6 and αTC-ΔPC2 Cell of the of expression of PC1/3 in the αTC-ΔPC2 cells, we a gene expression as M.S. C. Steiner D.F. 2001; 50: Scholar), to a of findings are in I. The the expression of PC1/3 low levels in αTC-ΔPC2 cells, but not in the αTC1–6 cell However, many other including of proglucagon, not The same for other neuroendocrine to both and such as and also for PC1/3 a low but of which with the observed of gene expression. of by both and analysis expression in both cell not of expression analysis of αTC1–6 and and in a The differential processing of proglucagon in pancreatic α cells to glucagon and in intestinal L cells to and has been a major of (28Rouille Y. Westermark G. Martin S.K. Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 3242-3246Google Scholar, 29Rouille Y. Martin S. Steiner D.F. J. Biol. Chem. 1995; 270: 26488-26496Google Scholar, 30Rouillé Y. Bianchi M. Irminger J.-C. Halban P. FEBS Lett. 1997; 413: 119-123Google Scholar, 31Rouillé Y. Kantengwa S. Irminger J.-C. Halban P.A. J. Biol. Chem. 1997; 272: Scholar). Although the PC family of convertases is and expressed in multiple the enzymes responsible for neuroendocrine proprotein processing are PC1/3 and PC2 (13Zhou A. Webb G. Zhu X. Steiner D.F. J. Biol. Chem. 1999; 274: 20745-20748Google Scholar). PC2 null in proglucagon processing in pancreatic only small amounts of glicentin and mature glucagon M. A. Y. R. M. L. Furuta Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1997; Scholar, M. A. Webb G. R. M. L. Steiner D.F. J. Biol. Chem. 2001; Scholar). This is with by a in α-cell in the Although PC2 is expressed in many other neuroendocrine cells in the that the of active glucagon about the has been in glucagon of glucagon in these led to of both levels and α-cell M.S. C. Steiner D.F. Scholar). However, of between cell expression of convertases in the cell and in α-cells from many the function of the PC2 null α-cells these we have derived an α-cell line from PC2 null that has the of glucagon processing in α-cells in the absence of PC2 activity. The characterized cells have been designated αTC-ΔPC2 cells. αTC-ΔPC2 cells the αTC1–6 cells with a and secretory granules with M. A. Y. R. M. L. Furuta Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1997; Scholar). that these cells express and for α-cells PC2 activity. expression analysis also provides a detailed analysis of cellular M.S. C. Steiner D.F. 2001; 50: Scholar), and findings the αTC-ΔPC2 and αTC1–6 cell many further with the of the of low levels of and PC1/3 and expression of to the characterized from PC2 null αTC-ΔPC2 cells a of PC2 that is not and activity and The expression of the PC2 and production of the for levels to that of PC2 in α-cells further the α-cell of these cells. the truncated PC2 is is of levels are not of proglucagon processing in αTC-ΔPC2 cell a complete of mature glucagon as PC2 is for or more of the required processing and The major is not proglucagon, but the of the cells to the cleavage the dibasic site amino acids of proglucagon processing in from PC2 null a more of the to M. A. Webb G. R. M. L. Steiner D.F. J. Biol. Chem. 2001; proteins from the αTC-ΔPC2 cells further that an additional is in these cells that is able to cleave the dibasic and to small amounts of and both the N-terminally extended of and GLP-2. additional the site to generate low amounts of and analysis that the αTC-ΔPC2 line but levels of PC1/3 with other cell that are known to express PC1/3 and This is further by analysis both α-cells and the αTC1–6 cell expression of PC1/3 is similar to cells and These findings that the initial processing the dibasic and the less processing the and sites in αTC-ΔPC2 cells be by the small amounts of PC1/3 in cell The of small amounts of truncated is also consistent with Y. Kantengwa S. Irminger J.-C. Halban P.A. J. Biol. Chem. 1997; 272: Scholar). PC1/3 is in the L cell processing of proglucagon to and is by recent studies on PC1/3 X. A. A. Norrbom C. R. Zhang C. Lindberg I. R. Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. Scholar). from these mainly proglucagon and only small amounts of glicentin X. A. A. Norrbom C. R. Zhang C. Lindberg I. R. Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. Scholar, R. Zhu X. C. Steiner D.F. Endocrinology. 2004; 145: Scholar). However, is in proglucagon processing in the of these and the of the is in to the PC2 null α-cells X. L. R. Norrbom C. M. Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. Scholar). of proglucagon processing in a variety of neuroendocrine cells levels of both PC1/3 PC2 that the site is to cleavage (28Rouille Y. Westermark G. Martin S.K. Steiner D.F. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 3242-3246Google Scholar, 29Rouille Y. Martin S. Steiner D.F. J. Biol. Chem. 1995; 270: 26488-26496Google Scholar, 30Rouillé Y. Bianchi M. Irminger J.-C. Halban P. FEBS Lett. 1997; 413: 119-123Google Scholar, 31Rouillé Y. Kantengwa S. Irminger J.-C. Halban P.A. J. Biol. Chem. 1997; 272: Scholar). is in both α and L cells, that both enzymes The low levels of PC1/3 in αTC-ΔPC2 cells, only low levels of the L cell including oxyntomodulin, GLP-1, extended and The of expression of neuroendocrine processing in the α-cell also the overall we the and characterization of the αTC-ΔPC2 cell a pancreatic α-cell line of PC2 activity. of these cells that α-cell is not on PC2 activity or the production of mature glucagon. The analysis that proglucagon be to mature glucagon in the absence of The cells, express low levels of which in a of proglucagon Y. M. R. Y. D. J. have expression of PC1/3 in α-cells of However, such of PC1/3 observed in the α-cells of PC2 null M. A. Webb G. R. M. L. Steiner D.F. J. Biol. Chem. 2001; Scholar). the low expression of PC1/3 and observed in these cells is an of their M. P. B. Mol. Cell. Endocrinol. 1999; 10: Scholar, M.E. M.S. Dev. Scholar, M. Y. M. Webb G. Furuta M. Steiner D. G. Endocrinology. 2003; is an We for on cell line for with and and for