from sso to ngs in hla population studies: the case of...

ContextTyping comparisonsPopulation Genetics

Conclusion

From SSO to NGS in HLA population studies:the case of Mandenka (Senegal)

Thomas Goeury, Lydie Brunet, Lisa Creary, Maeva Pasquier,Jean-Marie Tiercy, Maxime Galan, Barbara Kervaire,

Marcelo A. Fernandez Vinia, André Langaney, José Manuel Nunesand Alicia Sanchez-Mazas

Contact: [email protected]

11 May 2016

[EFI 2016 Population Genetics Working Group] T. Goeury HLA: From SSO to NGS


Conclusion

Materials & Methods

Genetic data & Typing methods

Genetic data

Mandenka (Niger-congo)Western Africa – Senegal

3 Class II HLA genes:HLA-DPB1 ; -DRB1 ; -DQB1

Typings

1 PCR-SSO (Geneva 1992)2 NGS:

Exon 2: 454 pyrosequencing(Geneva 2012)

Full gene: MiSeq(Stanford and Geneva, 2015-2016)

Mali

Figure : Sampling location in the Kedougou city

neighborhood, eastern Senegal



Conclusion

DifferencesCauses of differences

Comparisons of techniquesH

LA D

PB

1H

LA D

QB

1H

LA D

RB

1

Mea

n

HLA

DP

B1

HLA

DQ

B1

HLA

DR

B1

Mea

n

HLA

DP

B1

HLA

DQ

B1

HLA

DR

B1

Mea

n

454 versus MiSeq SSO versus 454 SSO versus MiSeq

0

25

50

75

100

Mat

ch p

ropo

rtio

ns

Proportions of typing matches between SSO, 454and MiSeq techniques for the 3 HLA Class II loci

Good match between NGS:DNA sequences

Correct match between454 & SSO:Exon 2

Discrepancies betweenSSO & MiSeq:

Locus-dependant



Conclusion


Years of allele reports

1980 1990 2000 2010

454MiSeq

NGS ERA

14:5401:01; 03:01; 03:02; 04:03; 04:05 07:01; 08:04; 08:06; 09:01; 10:0111:01; 11:02; 13:01; 13:02; 13:03

13:04; 16:02

SSO

NGS ERA

02:01; 02:02; 03:01; 03:0204:02; 05:01;05:02; 05:03

06:02; 06:03

06:09 03:1903:01:04

1980 1990 2000 2010NGS ERA

124:01; 131:01; 417:01430:01; 461:01

02:0103:0104:01

01:01; 10:0113:01; 17:01

01:01:02 85:01 104:01105:01107:01

1980 1990 2000 2010

HLA-DPB1HLA-DPB1

HLA-DQB1HLA-DQB1

HLA-DRB1HLA-DRB1



Conclusion


Resolutive power of Class II HLA exons

Exo

n 1

Exo

n 2

Exo

ns 2

&3

Exo

n 3

Exo

n 4

Exo

ns 5

&6

Exo

n 1

Exo

n 2

Exo

ns 2

&3

Exo

n 3

Exo

n 4

Exo

ns 5

&6

Exo

n 1

Exo

n 2

Exo

ns 2

&3

Exo

n 3

Exo

ns 4

&5

HLA−DQB1 HLA−DRB1 HLA−DPB1

−25

0

25

50

75

100

Percent of unique haplotypes (compared to max)per exon and per locus



Conclusion

Alleles FrequenciesLinkage disequilibriumsHLA-DRB1*13:04

HLA-DQB1: Frequencies in Mandenka

DQB1*03:02

DQB1*04:01

DQB1*06:05

DQB1*06:06

DQB1*05:03

DQB1*06:04

DQB1*04:02

DQB1*05:02

DQB1*06:03

DQB1*06:02

DQB1*05:01

DQB1*02

DQB1*03:01

0.0 0.1 0.2 0.3 0.4 0.5

Frequencyrange

>10%

5%−10%

2%−5%

1%−2%

10%

5%−10%

2%−5%

1%−2%


Conclusion


HLA-DPB1: Frequencies in Mandenka

DPB1*15:01

DPB1*39:01

DPB1*50:01

DPB1*11:01

DPB1*14:01

DPB1*40:01

DPB1*27:01

DPB1*03:01

DPB1*04:01

DPB1*13:01

DPB1*04:02

DPB1*01:01

DPB1*02:01

DPB1*17:01

0.0 0.1 0.2 0.3 0.4

Frequencyrange

>10%

5%−10%

2%−5%

1%−2%


Conclusion


HLA-DRB1: Frequencies in Mandenka

DRB1*01:01

DRB1*01:02

DRB1*04:03

DRB1*04:05

DRB1*09:01

DRB1*12:01

DRB1*08:01

DRB1*08:06

DRB1*14:01

DRB1*08:04

DRB1*16:01

DRB1*08:02

DRB1*13:03

DRB1*03:02

DRB1*13:01

DRB1*13:02

DRB1*10:01

DRB1*11:02

DRB1*07

DRB1*11:01

DRB1*03:01

DRB1*13:04

0.0 0.1 0.2 0.3

Frequencyrange

>10%

5%−10%

2%−5%

1%−2%

10%

5%−10%

2%−5%

1%−2%


Conclusion


Haplotypic and global linkage disequilibrium

Chr.6

Long

arm

Sho

rt a

rmC

ento

rom

ere

Tel

omer

eT

elom

ere

HCP5P15

PSMB9

Telomeric side

Centromeric side

HCG4P11HLA-FRPL23AP1MICEHCG9P5IFITM4P3.8-1.5HCP5P14HCG4P10HLA-75HCG4HCP5P13HLA-90

TUBB

LOC100129921

LSM2

BAT1

ATP6V1G2NFKBIL1LTATNFLTBLST1NCR3

AIF1BAT2

BAT3APOMC6orf47BAT4CSNK2BLY6G5BLY6G5CBAT5LY6G6FLY6G6ELY6G6D

C6orf25DDAH2CLIC1MSH5

C6orf27VARS

HSPA1LHSPA1AHSPA1BC6orf48

NEU1SLC44A4

EHMT2ZBTB12C2CFBRDBPSKIV2LDOM3ZSTK19C4BCYP21A1P

TNXBCREBL1FKBPLPRRT1PPT2EGFL8AGPAT1

HLA-DRAHLA-DRB9HLA-DRB5HLA-DRB6HLA-DRB1HLA-DQA1HLA-DQB1HLA-DQB3HLA-DQA2HLA-DQB2HLA-DOBTAP2PSMB8TAP1

PPP1R2P1HLA-ZHLA-DMBHLA-DMABRD2HLA-DOAHLA-DPA1

RPL32P1HLA-DPB1

HLA-DPA2HLA-DPB2HLA-DPA3COL11A2RXRBSLC39A7HSD17B8RING1VPS52RPS18B3GALT4

D6S2723EWDR46

PFDN6RGL2TAPBPZBTB22DAXXLOC646720

KIFC1LYPLA2P1

RPL7AP7MICGHCG2P8HCP5P12HCG4P8P5-11HLA-GLOC100133214

MICF3.8-1.4HCP5P10HCG4P7P5-09HLA-HP5-07HLA-16HCG2P73.8-1.3HCP5P6HCG4P6P5-05HLA-KHLA-UHCG4P5P5-04HLA-AHCP5P3HCG4P4HLA-WHCG2P6MICDHCG93.8-1.2HCP5P2HCG4P3HLA-J

ETF1P1

ZNRD1PPP1R11RNF39TRIM31TRIM40TRIM10

FLJ45422

TRIM39RPP21HLA-NHCG2P5HCG2P4MICCHCG2P3RANP1HLA-EGNL1PRR3ABCF1PPP1R10MRPS18BPTMAP1C6orf134C6orf136DHX16KIAA1949NRMRPL7APMDC1

FLOT1IER3DDR1

TCF19

GTF2H4VARS2

DPCR1MUC21

HCG22

CDSNPSORS1C1PSORS1C2

CCHCR1

POU5F1

HCG27HCG2P2HCG9P3HLA-CHCG4P2KIAA0055PRPL3PHCG2P1HLA-BHCG4P1DHFRP2HLA-SHCP5P8HCG9P2MICAHLA-XHCP53.8-1HCG9P1MICB

LOC100129065

HCG4P9

MCCD1

Cla

ss I

regi

on

Cla

ss I

regi

on

Cla

ss II

I reg

ion

Cla

ss II

reg

ion

Ext

ende

d cl

ass

II re

gion

Alp

h ab l

ock

Kap

p ab l

ock

Bet

abl

o ck

ZNFsORsGABBR1SUMO2PMOGZNP57

LOC100133303

SFTA2

LOC729792

C6orf15

LOC100129610

PSORS1C3

TRIM15

HLA-LPPIP9

SNORD117SNORD84

LOC100130756

SNORA38

LY6G6C

C6orf26

SNORD48SNORD52

LOC100128067

TNXASTK19PC4ACYP21A2

LOC100131609

RNF5AGERPBX2GPSM3NOTCH4C6orf10

BTNL2

MYL8P

HCG8

C6orf12

TRIM26

Figure 1 Gene map of the human leukocyte antigen (HLA) region. The major histocompatibility complex (MHC) gene map corresponds to the genomiccoordinates of 29677984 (GABBR1) to 33485635 (KIFC1) in the human genome build 36.3 of the National Center for Biotechnology Information (NCBI)map viewer. The regions separated by arrows show the HLA subregions such as extended class I, classical class I, class III, classical class II and extendedclass II regions from telomere (left and top side) to centromere (right and bottom side). White, gray, striped and black boxes show expressed genes, genecandidates, non-coding genes and pseudogenes, respectively. The location of the alpha, beta and kappa blocks containing the cluster of duplicated HLAclass I genes in the class I region are indicated.

A 2009 update of the HLA genomic loci mapT Shiina et al

24

Journal of Human Genetics

Figure : Adapted fromShiina et al, 2009

Haplotypes in Linkage Disequilibrium

DRB1*13:04 ∼ DQB1*03:19DRB1*11:02:01 ∼ DQB1*03:19

DRB1*03:01:01:01 ∼ DQB1*02:01:01–

Consequence of typing accuracy:DPB1*17:01-DQB1*03:01 no more in LD

(DPB1*17:01SSO = 17:01 + 131:01)

Global Linkage Disequilibrium between loci

HLA-DRB1 ∼ HLA-DQB1



Conclusion


Hypothesis of gene conversion

Hill et al (1992)

”The DRB1*1304 allele differs by only two amino acids, 57 and 58,from the DRB1*1102 allele, and it was suggested that a smallnonreciprocal genetic exchange event, perhaps with a DRB1*0803allele as the donor, led to the creation of the DRB1*1304 allele.”

AA Codon 55 60

DRB1*11:02:01 CGG CCT GAT GAG GAG TAC

DRB1*13:04 --- --- AGC -CC --- ---

Ser Ala



Conclusion


Gene conversion – New insigths

Donor and recipient

Recipient: DRB1*11:02:01

Donor ? Not DRB1*08:03

Presence of specific Ser-Ala pattern

Absent in this study, very rare in Africa

Two potentials donors: DRB1*08:06 & DRB1*13:03:01

Ser-Ala pattern, conserved region, present in Mandenka

Role ?



Conclusion

ConclusionAcknowledgement

Highlights

Key points

Importance of accurate typing using NGS and long rangesequencing

High frequency of DRB1*13:04 and DQB1*03:19, in LD

DQB1*03:19 may be involved in Onchocerciasis resistance

Loci HLA-DRB1 and HLA-DQB1 in strong LD

DRB1*13:04: hypothesis of a gene conversion betweenDRB1*11:02:01 and either DRB1*08:06 or DRB1*13:03:01



Conclusion


Acknowledgement

Εφςηαριστό για τιν προσοςηı́ σαςThank you for your attention

And also many thanks to. . .

The Mandenka people



Conclusion



ContextMaterials & Methods

Typing comparisonsDifferencesCauses of differences

Population GeneticsAlleles FrequenciesLinkage disequilibriumsHLA-DRB1*13:04

ConclusionConclusionAcknowledgement

from sso to ngs in hla population studies: the case of...

Documents