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The activation of IgM- or isotype-switched IgG- and IgE-BCR exhibits distinct mechanical force sensitivity and threshold.

Wan Z, Chen X, Chen H, Ji Q, Chen Y, Wang J, Cao Y, Wang F, Lou J, Tang Z, Liu W - Elife (2015)

Bottom Line: We observed that IgM-BCR activation is dependent on mechanical forces and exhibits a multi-threshold effect.Mechanistically, we found that the cytoplasmic tail of the IgG-BCR heavy chain is both required and sufficient to account for the low mechanical force threshold.These results defined the mechanical force sensitivity and threshold that are required to activate different isotyped BCRs.

View Article: PubMed Central - PubMed

Affiliation: MOE Key Laboratory of Protein Sciences, Tsinghua University, Beijing, China.

ABSTRACT
B lymphocytes use B cell receptors (BCRs) to sense the physical features of the antigens. However, the sensitivity and threshold for the activation of BCRs resulting from the stimulation by mechanical forces are unknown. Here, we addressed this question using a double-stranded DNA-based tension gauge tether system serving as a predefined mechanical force gauge ranging from 12 to 56 pN. We observed that IgM-BCR activation is dependent on mechanical forces and exhibits a multi-threshold effect. In contrast, the activation of isotype-switched IgG- or IgE-BCR only requires a low threshold of less than 12 pN, providing an explanation for their rapid activation in response to antigen stimulation. Mechanistically, we found that the cytoplasmic tail of the IgG-BCR heavy chain is both required and sufficient to account for the low mechanical force threshold. These results defined the mechanical force sensitivity and threshold that are required to activate different isotyped BCRs.

No MeSH data available.


Related in: MedlinePlus

Quantification of the accumulation of IgG-BCR in recognition of NP-TGT sensors at different surface density.(A–C) Quantification of the synaptic accumulation of IgG-BCRs in J558L cells expressing memory B1-8-IgG-BCR that were placed on coverslip coated with NP-TGT sensors at a density of 29.0 molecule/μm2 (A), 19.0 molecule/μm2 (B), and 0.3 molecule/μm2 (C). 12 pN NP-TGT, 56 pN NP-TGT, and 56 pN TGT molecule without NP conjugation (NC) were used in the experiment. Bars represent mean ±SEM. Two-tailed t tests were performed for the statistical comparisons. Data were from at least 30 cells over three independent experiments.DOI:http://dx.doi.org/10.7554/eLife.06925.015
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fig6s1: Quantification of the accumulation of IgG-BCR in recognition of NP-TGT sensors at different surface density.(A–C) Quantification of the synaptic accumulation of IgG-BCRs in J558L cells expressing memory B1-8-IgG-BCR that were placed on coverslip coated with NP-TGT sensors at a density of 29.0 molecule/μm2 (A), 19.0 molecule/μm2 (B), and 0.3 molecule/μm2 (C). 12 pN NP-TGT, 56 pN NP-TGT, and 56 pN TGT molecule without NP conjugation (NC) were used in the experiment. Bars represent mean ±SEM. Two-tailed t tests were performed for the statistical comparisons. Data were from at least 30 cells over three independent experiments.DOI:http://dx.doi.org/10.7554/eLife.06925.015

Mentions: B cells use different isotypes of BCRs to recognize antigens and to initiate transmembrane activation signaling. Mature naive B cells use IgM-BCRs (termed naive IgM-BCR thereafter), while memory B cells mainly use IgG-BCRs (termed memory IgG-BCR thereafter) with a fraction of these use IgE-BCRs (termed memory IgE-BCR thereafter). Memory B cells are responsible for the rapid antigen recall humoral responses upon vaccine immunization (McHeyzer-Williams and McHeyzer-Williams, 2005; Pierce and Liu, 2010). Here, we explore the sensitivities and thresholds toward mechanical forces in the activation of memory B cells expressing isotype-switched IgG-BCRs or IgE-BCRs. We addressed this question using J558L cells expressing memory B1-8-IgG-BCR in the same experimental system that has been used for the naive B1-8-IgM-BCR-expressing J558L cells as described in detail above. Surprisingly, we observed a totally different pattern in terms of the dependence on mechanical forces for the activation of memory IgG-BCRs. Each of the three NP-TGT sensors producing mechanical forces at 12, 43, or 56 pN similarly drove the cells to undergo spreading responses and the synaptic accumulation of BCRs or pSyk signaling molecules into the IS (Figure 6A–D). This suggested that the activation of memory IgG-BCRs requires either no tension or a mechanical force below the mean rupture force of the lowest-force NP-TGT we used (12 pN). Similar patterned activation of memory IgG-BCR was observed at different surface density of NP-TGT sensors on coverslip (Figure 6—figure supplement 1A–C). Furthermore, a similar phenomenon was observed when examining the activation of B1-8 primary B cells expressing memory IgG-BCRs that were derived from an in vitro class-switch response following our published protocol (Liu et al., 2010b) (Figure 6E,F). We also used high speed TIRFM imaging to test the dynamics of memory IgG-BCR accumulation into the B cell IS starting with the initial immune recognition of memory IgG-BCRs with 12 pN, 56 pN and NP non-conjugated control TGT molecules within a 13-min time course (Figure 6G,H, Video 3). It was clear that 12 pN and 56 pN NP-TGT molecules similarly drove the accumulation of BCRs into the B cell IS. All of these results show that the activation of the memory IgG-BCR is independent on mechanical forces ranging from 12 to 56 pN as tested in our NP-TGT experimental system.10.7554/eLife.06925.014Figure 6.The activation of isotype-switched IgG-BCRs or IgE-BCRs on memory B cells requires either no tension or a mechanical force below 12 pN.


The activation of IgM- or isotype-switched IgG- and IgE-BCR exhibits distinct mechanical force sensitivity and threshold.

Wan Z, Chen X, Chen H, Ji Q, Chen Y, Wang J, Cao Y, Wang F, Lou J, Tang Z, Liu W - Elife (2015)

Quantification of the accumulation of IgG-BCR in recognition of NP-TGT sensors at different surface density.(A–C) Quantification of the synaptic accumulation of IgG-BCRs in J558L cells expressing memory B1-8-IgG-BCR that were placed on coverslip coated with NP-TGT sensors at a density of 29.0 molecule/μm2 (A), 19.0 molecule/μm2 (B), and 0.3 molecule/μm2 (C). 12 pN NP-TGT, 56 pN NP-TGT, and 56 pN TGT molecule without NP conjugation (NC) were used in the experiment. Bars represent mean ±SEM. Two-tailed t tests were performed for the statistical comparisons. Data were from at least 30 cells over three independent experiments.DOI:http://dx.doi.org/10.7554/eLife.06925.015
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4555871&req=5

fig6s1: Quantification of the accumulation of IgG-BCR in recognition of NP-TGT sensors at different surface density.(A–C) Quantification of the synaptic accumulation of IgG-BCRs in J558L cells expressing memory B1-8-IgG-BCR that were placed on coverslip coated with NP-TGT sensors at a density of 29.0 molecule/μm2 (A), 19.0 molecule/μm2 (B), and 0.3 molecule/μm2 (C). 12 pN NP-TGT, 56 pN NP-TGT, and 56 pN TGT molecule without NP conjugation (NC) were used in the experiment. Bars represent mean ±SEM. Two-tailed t tests were performed for the statistical comparisons. Data were from at least 30 cells over three independent experiments.DOI:http://dx.doi.org/10.7554/eLife.06925.015
Mentions: B cells use different isotypes of BCRs to recognize antigens and to initiate transmembrane activation signaling. Mature naive B cells use IgM-BCRs (termed naive IgM-BCR thereafter), while memory B cells mainly use IgG-BCRs (termed memory IgG-BCR thereafter) with a fraction of these use IgE-BCRs (termed memory IgE-BCR thereafter). Memory B cells are responsible for the rapid antigen recall humoral responses upon vaccine immunization (McHeyzer-Williams and McHeyzer-Williams, 2005; Pierce and Liu, 2010). Here, we explore the sensitivities and thresholds toward mechanical forces in the activation of memory B cells expressing isotype-switched IgG-BCRs or IgE-BCRs. We addressed this question using J558L cells expressing memory B1-8-IgG-BCR in the same experimental system that has been used for the naive B1-8-IgM-BCR-expressing J558L cells as described in detail above. Surprisingly, we observed a totally different pattern in terms of the dependence on mechanical forces for the activation of memory IgG-BCRs. Each of the three NP-TGT sensors producing mechanical forces at 12, 43, or 56 pN similarly drove the cells to undergo spreading responses and the synaptic accumulation of BCRs or pSyk signaling molecules into the IS (Figure 6A–D). This suggested that the activation of memory IgG-BCRs requires either no tension or a mechanical force below the mean rupture force of the lowest-force NP-TGT we used (12 pN). Similar patterned activation of memory IgG-BCR was observed at different surface density of NP-TGT sensors on coverslip (Figure 6—figure supplement 1A–C). Furthermore, a similar phenomenon was observed when examining the activation of B1-8 primary B cells expressing memory IgG-BCRs that were derived from an in vitro class-switch response following our published protocol (Liu et al., 2010b) (Figure 6E,F). We also used high speed TIRFM imaging to test the dynamics of memory IgG-BCR accumulation into the B cell IS starting with the initial immune recognition of memory IgG-BCRs with 12 pN, 56 pN and NP non-conjugated control TGT molecules within a 13-min time course (Figure 6G,H, Video 3). It was clear that 12 pN and 56 pN NP-TGT molecules similarly drove the accumulation of BCRs into the B cell IS. All of these results show that the activation of the memory IgG-BCR is independent on mechanical forces ranging from 12 to 56 pN as tested in our NP-TGT experimental system.10.7554/eLife.06925.014Figure 6.The activation of isotype-switched IgG-BCRs or IgE-BCRs on memory B cells requires either no tension or a mechanical force below 12 pN.

Bottom Line: We observed that IgM-BCR activation is dependent on mechanical forces and exhibits a multi-threshold effect.Mechanistically, we found that the cytoplasmic tail of the IgG-BCR heavy chain is both required and sufficient to account for the low mechanical force threshold.These results defined the mechanical force sensitivity and threshold that are required to activate different isotyped BCRs.

View Article: PubMed Central - PubMed

Affiliation: MOE Key Laboratory of Protein Sciences, Tsinghua University, Beijing, China.

ABSTRACT
B lymphocytes use B cell receptors (BCRs) to sense the physical features of the antigens. However, the sensitivity and threshold for the activation of BCRs resulting from the stimulation by mechanical forces are unknown. Here, we addressed this question using a double-stranded DNA-based tension gauge tether system serving as a predefined mechanical force gauge ranging from 12 to 56 pN. We observed that IgM-BCR activation is dependent on mechanical forces and exhibits a multi-threshold effect. In contrast, the activation of isotype-switched IgG- or IgE-BCR only requires a low threshold of less than 12 pN, providing an explanation for their rapid activation in response to antigen stimulation. Mechanistically, we found that the cytoplasmic tail of the IgG-BCR heavy chain is both required and sufficient to account for the low mechanical force threshold. These results defined the mechanical force sensitivity and threshold that are required to activate different isotyped BCRs.

No MeSH data available.


Related in: MedlinePlus