An atlas of B-cell clonal distribution in the human body

Abstract

B-cell responses result in clonal expansion, and can occur in a variety of tissues. To define how B-cell clones are distributed in the body, we sequenced 933,427 B-cell clonal lineages and mapped them to eight different anatomic compartments in six human organ donors. We show that large B-cell clones partition into two broad networks-one spans the blood, bone marrow, spleen and lung, while the other is restricted to tissues within the gastrointestinal (GI) tract (jejunum, ileum and colon). Notably, GI tract clones display extensive sharing of sequence variants among different portions of the tract and have higher frequencies of somatic hypermutation, suggesting extensive and serial rounds of clonal expansion and selection. Our findings provide an anatomic atlas of B-cell clonal lineages, their properties and tissue connections. This resource serves as a foundation for studies of tissue-based immunity, including vaccine responses, infections, autoimmunity and cancer.

Figure 1. Diversity, Similarity and Networks of Large Clones

(a) Peripheral blood clones exhibit the highest sampled diversity. Diversity of clones with at least 20 unique sequence instances (C₂₀ clones) is plotted at different orders (Hill numbers) in different tissues in D207. At an order of 0, the diversity is the number of different clones. At orders >1, diversity is influenced more by the most abundant clones. (b) Tissues exhibit higher internal similarity than blood. Box plots represent the distribution of cosine similarity between all pairs of sequencing libraries within a tissue (see Methods). Similarity is assessed for C₂₀ clones in D207. Boxes represent the first and third quartiles bisected by the median. Whiskers represent the most extreme data excluding outliers, where outliers (dots) are data beyond the third or first quartile by a distance exceeding 1.5 times the inter-quartile interval. Higher cosine values correspond to greater sharing of large clones between replicate libraries from the same tissue. (c) Large clones form two major networks– one in blood-rich compartments (red tones) and one in the GI tract (blue tones). Shown are the cosine similarities of C₂₀ clones between tissue pairs in D181 and D207. Each wedge within the circle represents a tissue. Each arrow represents the level of overlap (cosine similarity) in clones from other tissues to the clones in that wedge. Longer arrows indicate more overlap between the tissues. PBL = peripheral blood; BM = bone marrow; SPL = spleen; MLN = mesenteric lymph node.

Figure 2. Tissue Distributions of Large Clones

(a) Global (found in 6-8 tissues); (b) Regional (3-5 tissues) and (c) Two-Tissue C₂₀ Clones. Each line is a clone. Each circle denotes membership of the clone in a particular tissue. The size of circle represents the total number of sequence instances the clones have in each tissue (depicted in legend). The portion of the circle that is colored represents the fraction of sequencing libraries from that tissue that contain at least one sequence of the clone (with at least two copies). The frequencies of each distribution type are indicated to the right of each clone line. Only the most frequent tissue distribution types (those that are present in at least 5% of a given tissue category in at least one of the two donors—D181 or D207) are shown. Tissues are colored as in Fig 1. lun = lung; jej = jejunum; col = colon; other abbreviations are as in Fig. 1.

Figure 3. Analysis of Sequence Variants in Clonal Lineages

(a) Multi-tiered clonal lineages exhibit diversification and sharing of sequence variants within and between tissues. Trees are rooted in the closest germline VH gene allele in the IMGT database (see Methods). Numbers indicate somatic mutations. Circles are colored according to the tissue distribution of the sequence variants. Circle sizes are proportional to sequence copy numbers. Black dots indicate inferred nodes. Each clone is identified by an unique identifier number in http://immunedb.com/tissue-atlas. (b) GI tract tissue clones exhibit extensive sharing of sequence variants. The median of the distribution of clumpiness (a metric of sequence sharing within clonal lineages, see Methods) is shown for all two-tissues pairs across all C₂₀ clones. (c) Sequence sharing distributions within clonal lineages that are found in different tissue pairs. Clones with peripheral blood (PBL) and another tissue were the least mixed, followed by clones mixing blood and GI tract tissues, then blood tissue clones and finally GI tract clones were the most mixed. SPL = spleen.

Figure 4. Somatic Hypermutation in Different Tissues

(a) Clones are more mutated in the tissues. Shown are percentages of clones that have average mutation frequencies of 1% or more. In all clones, only sequences from a specific tissue are counted (see Methods). Each column represents a separate tissue. Each dot represents an individual donor. (b) GI tract clones have right-shifted mutation frequency distributions compared to blood tissue clones in most donors. The average number of mutations per clone is plotted versus the percent of clones with that mutation level. Each line denotes a separate tissue. Segregation of mutations per clone to different tissues was accomplished as in Fig. 4a (see Methods). (c) Lineage tree heights of C₂₀ single-tissue clones in different tissues of D207. Only tissues with >100 single tissue clones are shown. The tree height is defined as the maximum distance of a sequence in the clone from the germline when considering only 4-fold redundant synonymous mutations (see Methods).