- Email: [email protected]
Peptide-dependent HLA-KIR-mediated regulation of NK cell function
Accepted Manuscript Editorial Peptide-dependent HLA-KIR-mediated regulation of NK cell function Barbara Rehermann PII: DOI: Reference:
S0168-8278(16)30194-5 http://dx.doi.org/10.1016/j.jhep.2016.05.008 JHEPAT 6104
To appear in:
Journal of Hepatology
Received Date: Accepted Date:
6 May 2016 9 May 2016
Please cite this article as: Rehermann, B., Peptide-dependent HLA-KIR-mediated regulation of NK cell function, Journal of Hepatology (2016), doi: http://dx.doi.org/10.1016/j.jhep.2016.05.008
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Peptide-dependent HLA-KIR-mediated regulation of NK cell function
Barbara Rehermann
Immunology Section, Liver Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, DHHS, Bethesda, MD, USA
Corresponding Author: Dr. Barbara Rehermann, Immunology Section, Liver Diseases Branch, NIDDK, National Institutes of Health, DHHS, 10 Center Drive, Bldg. 10, Rm. 9B16, Bethesda, MD 20892. Phone: 301-402-7144; Fax: 301-402-0491; E-mail: [email protected]
Page 1 of 13
The human leukocyte antigen (HLA) gene complex is located on the short arm of chromosome 6 and represents the most polymorphic region of the human genome. The extraordinary diversity of HLA alleles is thought to be driven by the evolutionary pressure of the pathogens that humans (and vertebrates in general) encounter in different parts of the world [1]. HLA class I (HLA-A, B and C) molecules bind endogenously processed peptides in the endoplasmatic reticulum (ER), transport them to the cell surface and present them to CD8 T cells, thereby allowing immune cells to scan the cell’s antigenic content. In some instances, HLA class I molecules can also present peptides that are derived from external antigens (cross-presentation). In contrast, HLA class II (DP, DQ, DR) - bound peptides are exclusively derived from externally acquired antigens and presented to CD4 T cells.
As soon as hepatitis C virus (HCV) starts protein synthesis in an infected cell it changes the repertoire of HLA-bound peptides with many self peptides being replaced by viral peptides. HCV also increases expression of antigen-processing enzymes such as the immunoproteasomes [2] and ER amino peptidases early after infection [3]. Because the magnitude, breadth and maintenance of the ultimate antiviral T cell responses correlates well with the outcome of infection [4] increased processing and presentation of antigenic peptides should not be in the interest of virus. Indeed, many other viruses have developed sophisticated strategies to interfere with this process [5] However, HCV-specific T cells take close to two months to appear in the circulation and ultimately in the liver [6]. Natural killer (NK) cells may pose an earlier thread to HCV because they constitute a large proportion of liver-resident immune cells and can exert
Page 2 of 13
immediate cytotoxic and cytokine-mediated effector functions. In this issue of The Journal of Hepatology, Lunemann et al. [7] identify a HLA-class I-presented HCV core peptide that increases the binding of HLA class I to an inhibitory NK cell receptor and downregulates NK cell function.
NK cells integrate signals from a large number of activating and inhibitory receptors and are also stimulated by virus-induced cytokines. An important class of inhibitory receptors are killer immunoglobulin-like receptors (KIRs). They are expressed in a variegated manner on CD56dim NK cells (which constitute about 90% of the NK cell population), bind to HLA-A, B and C molecules and signal through long intraplasmatic tails with two immunoreceptor tyrosine-based inhibition motifs (ITIMs). Their activatory counterparts have short cytoplasmic tails with immunoreceptor tyrosine-based activation motifs (ITAMs). In addition, there are other classes of receptors [8]. In the absence of infection, inhibitory HLA-KIR signals dominate and protect cells from NK cell-mediated lysis (Fig. 1A). In contrast, the downregulation of HLA molecules (Fig. 2B) or the induction of cytokines such as type I IFN, IL-12, IL-15 and IL-18 by viruses can activate NK cells. How readily NK cells can get activated is to some extent genetically determined: Khakoo et al. described in a landmark publication that homozygosity for HLA-C group 1 alleles and KIR2DL3 predisposes a subgroup of patients to HCV clearance [9]. The HLA-C group 1 family comprises several HLA-C alleles (including the HLA-C*03:04 allele studied by Lunemann et al [7]) that share a serine at position 77 and asparagine at position 80, and are ligands to KIR2DL3. Homozygosity for HLA-C group 1 and KIR2DL3 is thought to result in relatively weaker KIR binding and signaling (compared to other HLA-KIR compound
Page 3 of 13
genotypes) [10], which would allow KIR2DL3+ NK cells from HLA-C group 1-positive subjects to respond more readily to signals from activatory receptors and inflammatory cytokines. KIR2DL3+ NK cells have also recently been shown to be more sensitive to changes in the peptide content of the HLA class I binding groove than NK cells with other KIRs [11]
Here, Lunemann et al. show that an HLA-C*03:04-restricted HCV core peptide with a genotype 1-specific sequence increases the binding between an HLA-C*03:04 and KIR2DL3, therefore downregulating the effector function of KIR2DL3+ NK cells.
The
authors start their investigation by screening 200 overlapping HCV core and NS3 peptides for binding to a B-cell line that does not express any HLA molecules other than HLAC*03:04. In this cell line, the loading of endogenous peptides onto HLA molecules is disabled by two experimental strategies. In one strategy the transporter associated with antigen processing (TAP) is deleted using CRISPR/CAS technology. In an alternative strategy the cells are transfected with a gene from a virus that inhibits the function of TAP. The absence of endogenously processed, high-affinity peptides increases the turnover of HLA molecules and reduces their expression on the cell surface.
This provides the
opportunity to pulse the cells with candidate peptides and to identify strong binders by their capacity to stabilize HLA-C*03:04 expression. The authors identify ten 15mer peptides as HLA-C*03:04 binders and identify the optimal binding sequences (9 mers) for two of these: the HCVcore136-144 peptide YIPLVGAPL and the HCV NS31325-1333 peptide SILGIGTVL. The HCVcore136-144 peptide and its longer 15mer precursor increase binding of an KIR2DL3IgG fusion construct to HLA-C*03:04. This increased engagement of KIR2DL3 results in
Page 4 of 13
reduced degranulation of KIR2DL3+ NK cells but not of KIR2DL3- NK cells from healthy blood donors.
Interestingly, the effect is specific for the genotype 1 sequence of the
core136-144 peptide and naturally occurring variants from other genotypes exhibit a reduced capacity to bind KIR2DL3 and to inhibit the function of KIR2DL3+ NK cells. These results imply that sequence variations in HCV may modulate NK cell function by modifying HLA/KIR interaction.
While the concept of peptides modulating the interaction of specific KIRs and their HLA-A, B or C ligands is not new [12-15] Lunemann et al. perform the first systematic screen for such peptides in HCV infection.
Only very few HLA-C restricted peptides have been
identified in HCV infection, even when one uses T cell activation rather than NK cell inhibition as a readout. A few points from the study should be emphasized: First, the authors demonstrate the specificity of the results by showing that the NK cell inhibitory effect depends on the HCV genotype 1 sequence of the core peptide, and that it is not observed when non-genotype 1 variants are used.
Second, they show that only
KIR2DL3+ and not KIRL2DL3- NK cells are inhibited. Third, they identify an HCV-infected patient with CD8 T cell responses against the core peptide. This data demonstrates that the core peptide is endogenously processed in HCV-infected humans and that it is able to induce peptide-specific T cells.
Of note, however, the inhibitory effect of HLA-
C*03:04+/HCV core peptide pulsed target cells on NK cells in vitro assays requires a 1001000-fold higher peptide concentration than typically required for T cell stimulation. This is in line with other studies [16]. While the inhibitory effect on NK cells is detectable at 10
Page 5 of 13
µΜ peptide concentration [7], T cell stimulation can often still be observed in the 0.01 to 0.1 µM range. Thus, the in vivo relevance of the findings requires further investigation.
An interesting step in this direction would be the screening of HCV sequences for amino acid polymorphisms that occur in the presence of specific HLA/KIR compound genotypes and specific outcomes of infection. Do the naturally occurring variant peptides that do not enhance KIR2DL3 binding act as antagonists to the HCVcore136-144 peptide YIPLVGAPL? Altered peptide ligands that do not have a high binding affinity and that, by themselves, do not inhibit KIR+ NK cells, may nevertheless antagonize the function of a high affinity peptide [17]. In a peptide mix, even low concentrations of such antagonist peptide can prevent the formation of KIR microclusters in response to the high-affinity peptide and thereby stall downstream signaling [17] (Fig. 1C). For multiple HLA-KIR combinations, amino acid positions 8 of the peptide has been described to be critical for HLA/KIR binding [12-14], and this has been confirmed by crystallization studies [18].
The design of the study and the lack of a cohort of HCV-infected patients did not allow the direct assessment of NK cell selection pressure during HCV sequence evolution. However, such a study was performed in an HIV cohort by some of the same authors [19]. This study identified more than 20 positions in the HIV-1 genome at which amino-acid polymorphisms were significantly associated with the presence of a specific KIR gene [19]. More recently, the same group confirmed that HIV escape mutations also occur in the context of the specific HLA-C*03:04/KIR2DLR3 compound genotype [20] that is now studied by Lunemann et al. for HCV. Collectively, these findings suggests that KIR-positive
Page 6 of 13
NK cells can indeed exert immunological pressure – a hypothesis that can be further explored with rapidly mutating viruses such as HIV and HCV.
Acknowledgement This work was supported by the intramural research program of NIDDK, NIH.
Page 7 of 13
References
[1]
Sommer S. The importance of immune gene variability (MHC) in evolutionary
ecology and conservation. Front Zool 2005;2:16. [2]
Shin EC, Seifert U, Kato T, Rice CM, Feinstone SM, Kloetzel PM, et al. Virus-
induced type I IFN stimulates generation of immunoproteasomes at the site of infection. J Clin Invest 2006;116:3006-3014. [3]
Shin EC, Seifert U, Urban S, Truong KT, Feinstone SM, Rice CM, et al. Proteasome
activator and antigen-processing aminopeptidases are regulated by virus-induced type I interferon in the hepatitis C virus-infected liver. J Interferon Cytokine Res 2007;27:985990. [4]
Rehermann B. Pathogenesis of chronic viral hepatitis: differential roles of T cells
and NK cells. Nat Med 2013;19:859-868. [5]
Petersen JL, Morris CR, Solheim JC. Virus evasion of MHC class I molecule
presentation. J Immunol 2003;171:4473-4478. [6]
Shin EC, Park SH, Demino M, Nascimbeni M, Mihalik K, Major M, et al. Delayed
induction, not impaired recruitment, of specific CD8(+) T cells causes the late onset of acute hepatitis C. Gastroenterology 2011;141:686-695, 695 e681. [7]
Lunemann S, Martrus G, Holzemer A, Chapel A, Ziegler M, Korner C, et al.
Sequence variations in HCV core-derived epitopes alter binding of KIR2DL3 to HLAC03:04 and modulate NK cell function. J Hepatol 2016. [8]
Lanier LL. Up on the tightrope: natural killer cell activation and inhibition. Nat
Immunol 2008;9:495-502.
Page 8 of 13
[9]
Khakoo SI, Thio CL, Martin MP, Brooks CR, Gao X, Astemborski J, et al. HLA and
NK cell inhibitory receptor genes in resolving hepatitis C virus infection. Science 2004;305:872-874. [10]
Winter CC, Gumperz JE, Parham P, Long EO, Wagtmann N. Direct binding and
functional transfer of NK cell inhibitory receptors reveal novel patterns of HLA-C allotype recognition. J Immunol 1998;161:571-577. [11]
Cassidy S, Mukherjee S, Myint TM, Mbiribindi B, North H, Traherne J, et al. Peptide
selectivity discriminates NK cells from KIR2DL2- and KIR2DL3-positive individuals. Eur J Immunol 2015;45:492-500. [12]
Rajagopalan S, Long EO. The direct binding of a p58 killer cell inhibitory receptor to
human histocompatibility leukocyte antigen (HLA)-Cw4 exhibits peptide selectivity. J Exp Med 1997;185:1523-1528. [13]
Hansasuta P, Dong T, Thananchai H, Weekes M, Willberg C, Aldemir H, et al.
Recognition of HLA-A3 and HLA-A11 by KIR3DL2 is peptide-specific. Eur J Immunol 2004;34:1673-1679. [14]
Thananchai H, Gillespie G, Martin MP, Bashirova A, Yawata N, Yawata M, et al.
Cutting Edge: Allele-specific and peptide-dependent interactions between KIR3DL1 and HLA-A and HLA-B. J Immunol 2007;178:33-37. [15]
Cassidy SA, Cheent KS, Khakoo SI. Effects of Peptide on NK cell-mediated MHC I
recognition. Front Immunol 2014;5:133. [16]
Fadda L, Borhis G, Ahmed P, Cheent K, Pageon SV, Cazaly A, et al. Peptide
antagonism as a mechanism for NK cell activation. Proc Natl Acad Sci U S A 2010;107:10160-10165.
Page 9 of 13
[17]
Borhis G, Ahmed PS, Mbiribindi B, Naiyer MM, Davis DM, Purbhoo MA, et al. A
peptide
antagonist
disrupts
NK
cell
inhibitory
synapse
formation.
J
Immunol
2013;190:2924-2930. [18]
Boyington JC, Motyka SA, Schuck P, Brooks AG, Sun PD. Crystal structure of an
NK cell immunoglobulin-like receptor in complex with its class I MHC ligand. Nature 2000;405:537-543. [19]
Alter G, Heckerman D, Schneidewind A, Fadda L, Kadie CM, Carlson JM, et al.
HIV-1 adaptation to NK-cell-mediated immune pressure. Nature 2011;476:96-100. [20]
Holzemer A, Thobakgale CF, Jimenez Cruz CA, Garcia-Beltran WF, Carlson JM,
van Teijlingen NH, et al. Selection of an HLA-C*03:04-Restricted HIV-1 p24 Gag Sequence Variant Is Associated with Viral Escape from KIR2DL3+ Natural Killer Cells: Data from an Observational Cohort in South Africa. PLoS Med 2015;12:e1001900; discussion e1001900.
Page 10 of 13
Figure 1: Simplified model detailing the regulation of NK cell activation by HLA-KIR interaction Peptide-loaded HLA class I complexes on target cells bind to killer immunoglobulin-like receptors (KIRs) of NK cells. Peptides are endogenously processed from longer precursors by the cytosolic proteasome and loaded into the endoplasmic reticulum by transporters associated with antigen processing (TAP). After N-terminal trimming by ER aminopeptidases (ERAAP), they bind to HLA class I molecules and are transported via the Golgi and secretory vesicles onto the cell surface. A. NK cells integrate signals from a large number of activating (shown in green) and inhibitory receptors (shown in red) and are also stimulated by virus-induced cytokines (not shown). KIRs are an important group of inhibitory receptors that bind to peptidepresenting HLA class I molecules (blue). In the presence of high affinity ligands they form clusters. In the healthy host, negative (-) signals dominate over positive (+) signals resulting in NK cell inhibition. B. The downregulation of HLA molecules (“missing self”) activates NK cells. The release of perforin/granzyme-containing granules results in NK cell cytotoxicity. The merging of the CD107a+ granules with the cell surface membrane increases CD107a levels on the cell surface, which can be assessed by flow cytometry as a read-out for NK cell degranulation. Induction of FasL and TRAIL can also contribute to cytotoxicity. Cytotoxicity and cytokine release can have divergent effects on virus-infected cells, antigen-presenting cells and T cells [4].
Page 11 of 13
C. Low concentrations of variant peptide (black) can prevent the formation of KIR microclusters in response to the high-affinity peptide (pink) and stall downstream signaling.
Page 12 of 13
Page 13 of 13
S0168-8278(16)30194-5 http://dx.doi.org/10.1016/j.jhep.2016.05.008 JHEPAT 6104
To appear in:
Journal of Hepatology
Received Date: Accepted Date:
6 May 2016 9 May 2016
Please cite this article as: Rehermann, B., Peptide-dependent HLA-KIR-mediated regulation of NK cell function, Journal of Hepatology (2016), doi: http://dx.doi.org/10.1016/j.jhep.2016.05.008
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.
Peptide-dependent HLA-KIR-mediated regulation of NK cell function
Barbara Rehermann
Immunology Section, Liver Diseases Branch, National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, DHHS, Bethesda, MD, USA
Corresponding Author: Dr. Barbara Rehermann, Immunology Section, Liver Diseases Branch, NIDDK, National Institutes of Health, DHHS, 10 Center Drive, Bldg. 10, Rm. 9B16, Bethesda, MD 20892. Phone: 301-402-7144; Fax: 301-402-0491; E-mail: [email protected]
Page 1 of 13
The human leukocyte antigen (HLA) gene complex is located on the short arm of chromosome 6 and represents the most polymorphic region of the human genome. The extraordinary diversity of HLA alleles is thought to be driven by the evolutionary pressure of the pathogens that humans (and vertebrates in general) encounter in different parts of the world [1]. HLA class I (HLA-A, B and C) molecules bind endogenously processed peptides in the endoplasmatic reticulum (ER), transport them to the cell surface and present them to CD8 T cells, thereby allowing immune cells to scan the cell’s antigenic content. In some instances, HLA class I molecules can also present peptides that are derived from external antigens (cross-presentation). In contrast, HLA class II (DP, DQ, DR) - bound peptides are exclusively derived from externally acquired antigens and presented to CD4 T cells.
As soon as hepatitis C virus (HCV) starts protein synthesis in an infected cell it changes the repertoire of HLA-bound peptides with many self peptides being replaced by viral peptides. HCV also increases expression of antigen-processing enzymes such as the immunoproteasomes [2] and ER amino peptidases early after infection [3]. Because the magnitude, breadth and maintenance of the ultimate antiviral T cell responses correlates well with the outcome of infection [4] increased processing and presentation of antigenic peptides should not be in the interest of virus. Indeed, many other viruses have developed sophisticated strategies to interfere with this process [5] However, HCV-specific T cells take close to two months to appear in the circulation and ultimately in the liver [6]. Natural killer (NK) cells may pose an earlier thread to HCV because they constitute a large proportion of liver-resident immune cells and can exert
Page 2 of 13
immediate cytotoxic and cytokine-mediated effector functions. In this issue of The Journal of Hepatology, Lunemann et al. [7] identify a HLA-class I-presented HCV core peptide that increases the binding of HLA class I to an inhibitory NK cell receptor and downregulates NK cell function.
NK cells integrate signals from a large number of activating and inhibitory receptors and are also stimulated by virus-induced cytokines. An important class of inhibitory receptors are killer immunoglobulin-like receptors (KIRs). They are expressed in a variegated manner on CD56dim NK cells (which constitute about 90% of the NK cell population), bind to HLA-A, B and C molecules and signal through long intraplasmatic tails with two immunoreceptor tyrosine-based inhibition motifs (ITIMs). Their activatory counterparts have short cytoplasmic tails with immunoreceptor tyrosine-based activation motifs (ITAMs). In addition, there are other classes of receptors [8]. In the absence of infection, inhibitory HLA-KIR signals dominate and protect cells from NK cell-mediated lysis (Fig. 1A). In contrast, the downregulation of HLA molecules (Fig. 2B) or the induction of cytokines such as type I IFN, IL-12, IL-15 and IL-18 by viruses can activate NK cells. How readily NK cells can get activated is to some extent genetically determined: Khakoo et al. described in a landmark publication that homozygosity for HLA-C group 1 alleles and KIR2DL3 predisposes a subgroup of patients to HCV clearance [9]. The HLA-C group 1 family comprises several HLA-C alleles (including the HLA-C*03:04 allele studied by Lunemann et al [7]) that share a serine at position 77 and asparagine at position 80, and are ligands to KIR2DL3. Homozygosity for HLA-C group 1 and KIR2DL3 is thought to result in relatively weaker KIR binding and signaling (compared to other HLA-KIR compound
Page 3 of 13
genotypes) [10], which would allow KIR2DL3+ NK cells from HLA-C group 1-positive subjects to respond more readily to signals from activatory receptors and inflammatory cytokines. KIR2DL3+ NK cells have also recently been shown to be more sensitive to changes in the peptide content of the HLA class I binding groove than NK cells with other KIRs [11]
Here, Lunemann et al. show that an HLA-C*03:04-restricted HCV core peptide with a genotype 1-specific sequence increases the binding between an HLA-C*03:04 and KIR2DL3, therefore downregulating the effector function of KIR2DL3+ NK cells.
The
authors start their investigation by screening 200 overlapping HCV core and NS3 peptides for binding to a B-cell line that does not express any HLA molecules other than HLAC*03:04. In this cell line, the loading of endogenous peptides onto HLA molecules is disabled by two experimental strategies. In one strategy the transporter associated with antigen processing (TAP) is deleted using CRISPR/CAS technology. In an alternative strategy the cells are transfected with a gene from a virus that inhibits the function of TAP. The absence of endogenously processed, high-affinity peptides increases the turnover of HLA molecules and reduces their expression on the cell surface.
This provides the
opportunity to pulse the cells with candidate peptides and to identify strong binders by their capacity to stabilize HLA-C*03:04 expression. The authors identify ten 15mer peptides as HLA-C*03:04 binders and identify the optimal binding sequences (9 mers) for two of these: the HCVcore136-144 peptide YIPLVGAPL and the HCV NS31325-1333 peptide SILGIGTVL. The HCVcore136-144 peptide and its longer 15mer precursor increase binding of an KIR2DL3IgG fusion construct to HLA-C*03:04. This increased engagement of KIR2DL3 results in
Page 4 of 13
reduced degranulation of KIR2DL3+ NK cells but not of KIR2DL3- NK cells from healthy blood donors.
Interestingly, the effect is specific for the genotype 1 sequence of the
core136-144 peptide and naturally occurring variants from other genotypes exhibit a reduced capacity to bind KIR2DL3 and to inhibit the function of KIR2DL3+ NK cells. These results imply that sequence variations in HCV may modulate NK cell function by modifying HLA/KIR interaction.
While the concept of peptides modulating the interaction of specific KIRs and their HLA-A, B or C ligands is not new [12-15] Lunemann et al. perform the first systematic screen for such peptides in HCV infection.
Only very few HLA-C restricted peptides have been
identified in HCV infection, even when one uses T cell activation rather than NK cell inhibition as a readout. A few points from the study should be emphasized: First, the authors demonstrate the specificity of the results by showing that the NK cell inhibitory effect depends on the HCV genotype 1 sequence of the core peptide, and that it is not observed when non-genotype 1 variants are used.
Second, they show that only
KIR2DL3+ and not KIRL2DL3- NK cells are inhibited. Third, they identify an HCV-infected patient with CD8 T cell responses against the core peptide. This data demonstrates that the core peptide is endogenously processed in HCV-infected humans and that it is able to induce peptide-specific T cells.
Of note, however, the inhibitory effect of HLA-
C*03:04+/HCV core peptide pulsed target cells on NK cells in vitro assays requires a 1001000-fold higher peptide concentration than typically required for T cell stimulation. This is in line with other studies [16]. While the inhibitory effect on NK cells is detectable at 10
Page 5 of 13
µΜ peptide concentration [7], T cell stimulation can often still be observed in the 0.01 to 0.1 µM range. Thus, the in vivo relevance of the findings requires further investigation.
An interesting step in this direction would be the screening of HCV sequences for amino acid polymorphisms that occur in the presence of specific HLA/KIR compound genotypes and specific outcomes of infection. Do the naturally occurring variant peptides that do not enhance KIR2DL3 binding act as antagonists to the HCVcore136-144 peptide YIPLVGAPL? Altered peptide ligands that do not have a high binding affinity and that, by themselves, do not inhibit KIR+ NK cells, may nevertheless antagonize the function of a high affinity peptide [17]. In a peptide mix, even low concentrations of such antagonist peptide can prevent the formation of KIR microclusters in response to the high-affinity peptide and thereby stall downstream signaling [17] (Fig. 1C). For multiple HLA-KIR combinations, amino acid positions 8 of the peptide has been described to be critical for HLA/KIR binding [12-14], and this has been confirmed by crystallization studies [18].
The design of the study and the lack of a cohort of HCV-infected patients did not allow the direct assessment of NK cell selection pressure during HCV sequence evolution. However, such a study was performed in an HIV cohort by some of the same authors [19]. This study identified more than 20 positions in the HIV-1 genome at which amino-acid polymorphisms were significantly associated with the presence of a specific KIR gene [19]. More recently, the same group confirmed that HIV escape mutations also occur in the context of the specific HLA-C*03:04/KIR2DLR3 compound genotype [20] that is now studied by Lunemann et al. for HCV. Collectively, these findings suggests that KIR-positive
Page 6 of 13
NK cells can indeed exert immunological pressure – a hypothesis that can be further explored with rapidly mutating viruses such as HIV and HCV.
Acknowledgement This work was supported by the intramural research program of NIDDK, NIH.
Page 7 of 13
References
[1]
Sommer S. The importance of immune gene variability (MHC) in evolutionary
ecology and conservation. Front Zool 2005;2:16. [2]
Shin EC, Seifert U, Kato T, Rice CM, Feinstone SM, Kloetzel PM, et al. Virus-
induced type I IFN stimulates generation of immunoproteasomes at the site of infection. J Clin Invest 2006;116:3006-3014. [3]
Shin EC, Seifert U, Urban S, Truong KT, Feinstone SM, Rice CM, et al. Proteasome
activator and antigen-processing aminopeptidases are regulated by virus-induced type I interferon in the hepatitis C virus-infected liver. J Interferon Cytokine Res 2007;27:985990. [4]
Rehermann B. Pathogenesis of chronic viral hepatitis: differential roles of T cells
and NK cells. Nat Med 2013;19:859-868. [5]
Petersen JL, Morris CR, Solheim JC. Virus evasion of MHC class I molecule
presentation. J Immunol 2003;171:4473-4478. [6]
Shin EC, Park SH, Demino M, Nascimbeni M, Mihalik K, Major M, et al. Delayed
induction, not impaired recruitment, of specific CD8(+) T cells causes the late onset of acute hepatitis C. Gastroenterology 2011;141:686-695, 695 e681. [7]
Lunemann S, Martrus G, Holzemer A, Chapel A, Ziegler M, Korner C, et al.
Sequence variations in HCV core-derived epitopes alter binding of KIR2DL3 to HLAC03:04 and modulate NK cell function. J Hepatol 2016. [8]
Lanier LL. Up on the tightrope: natural killer cell activation and inhibition. Nat
Immunol 2008;9:495-502.
Page 8 of 13
[9]
Khakoo SI, Thio CL, Martin MP, Brooks CR, Gao X, Astemborski J, et al. HLA and
NK cell inhibitory receptor genes in resolving hepatitis C virus infection. Science 2004;305:872-874. [10]
Winter CC, Gumperz JE, Parham P, Long EO, Wagtmann N. Direct binding and
functional transfer of NK cell inhibitory receptors reveal novel patterns of HLA-C allotype recognition. J Immunol 1998;161:571-577. [11]
Cassidy S, Mukherjee S, Myint TM, Mbiribindi B, North H, Traherne J, et al. Peptide
selectivity discriminates NK cells from KIR2DL2- and KIR2DL3-positive individuals. Eur J Immunol 2015;45:492-500. [12]
Rajagopalan S, Long EO. The direct binding of a p58 killer cell inhibitory receptor to
human histocompatibility leukocyte antigen (HLA)-Cw4 exhibits peptide selectivity. J Exp Med 1997;185:1523-1528. [13]
Hansasuta P, Dong T, Thananchai H, Weekes M, Willberg C, Aldemir H, et al.
Recognition of HLA-A3 and HLA-A11 by KIR3DL2 is peptide-specific. Eur J Immunol 2004;34:1673-1679. [14]
Thananchai H, Gillespie G, Martin MP, Bashirova A, Yawata N, Yawata M, et al.
Cutting Edge: Allele-specific and peptide-dependent interactions between KIR3DL1 and HLA-A and HLA-B. J Immunol 2007;178:33-37. [15]
Cassidy SA, Cheent KS, Khakoo SI. Effects of Peptide on NK cell-mediated MHC I
recognition. Front Immunol 2014;5:133. [16]
Fadda L, Borhis G, Ahmed P, Cheent K, Pageon SV, Cazaly A, et al. Peptide
antagonism as a mechanism for NK cell activation. Proc Natl Acad Sci U S A 2010;107:10160-10165.
Page 9 of 13
[17]
Borhis G, Ahmed PS, Mbiribindi B, Naiyer MM, Davis DM, Purbhoo MA, et al. A
peptide
antagonist
disrupts
NK
cell
inhibitory
synapse
formation.
J
Immunol
2013;190:2924-2930. [18]
Boyington JC, Motyka SA, Schuck P, Brooks AG, Sun PD. Crystal structure of an
NK cell immunoglobulin-like receptor in complex with its class I MHC ligand. Nature 2000;405:537-543. [19]
Alter G, Heckerman D, Schneidewind A, Fadda L, Kadie CM, Carlson JM, et al.
HIV-1 adaptation to NK-cell-mediated immune pressure. Nature 2011;476:96-100. [20]
Holzemer A, Thobakgale CF, Jimenez Cruz CA, Garcia-Beltran WF, Carlson JM,
van Teijlingen NH, et al. Selection of an HLA-C*03:04-Restricted HIV-1 p24 Gag Sequence Variant Is Associated with Viral Escape from KIR2DL3+ Natural Killer Cells: Data from an Observational Cohort in South Africa. PLoS Med 2015;12:e1001900; discussion e1001900.
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Figure 1: Simplified model detailing the regulation of NK cell activation by HLA-KIR interaction Peptide-loaded HLA class I complexes on target cells bind to killer immunoglobulin-like receptors (KIRs) of NK cells. Peptides are endogenously processed from longer precursors by the cytosolic proteasome and loaded into the endoplasmic reticulum by transporters associated with antigen processing (TAP). After N-terminal trimming by ER aminopeptidases (ERAAP), they bind to HLA class I molecules and are transported via the Golgi and secretory vesicles onto the cell surface. A. NK cells integrate signals from a large number of activating (shown in green) and inhibitory receptors (shown in red) and are also stimulated by virus-induced cytokines (not shown). KIRs are an important group of inhibitory receptors that bind to peptidepresenting HLA class I molecules (blue). In the presence of high affinity ligands they form clusters. In the healthy host, negative (-) signals dominate over positive (+) signals resulting in NK cell inhibition. B. The downregulation of HLA molecules (“missing self”) activates NK cells. The release of perforin/granzyme-containing granules results in NK cell cytotoxicity. The merging of the CD107a+ granules with the cell surface membrane increases CD107a levels on the cell surface, which can be assessed by flow cytometry as a read-out for NK cell degranulation. Induction of FasL and TRAIL can also contribute to cytotoxicity. Cytotoxicity and cytokine release can have divergent effects on virus-infected cells, antigen-presenting cells and T cells [4].
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C. Low concentrations of variant peptide (black) can prevent the formation of KIR microclusters in response to the high-affinity peptide (pink) and stall downstream signaling.
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