L-selectin shedding is activated specifically within transmigrating pseudopods of monocytes to regulate cell polarity in vitro

Karolina Rzeniewicz; Abigail Newe; Angela Rey Gallardo; Jessic Daviesa; Mark R. Holt; Ashish Patel; Guillaume T. Charras; Brian Stramer; Chris Molenaar; Thomas F. Tedder; Maddy Parsons; Aleksandar Ivetic

doi:10.1073/pnas.1417100112

L-selectin shedding is activated specifically within transmigrating pseudopods of monocytes to regulate cell polarity in vitro

Karolina Rzeniewicz, Abigail Newe, Angela Rey Gallardo, Jessic Daviesa, Mark R. Holt, Ashish Patel, Guillaume T. Charras, Brian Stramer, Chris Molenaar, Thomas F. Tedder, Maddy Parsons, Aleksandar Ivetic^*

^*Corresponding author for this work

Research output: Contribution to journal › Article › peer-review

47 Citations (Scopus)

251 Downloads (Pure)

Abstract

L-selectin is a cell adhesion molecule that tethers free-flowing leukocytes from the blood to luminal vessel walls, facilitating the initial stages of their emigration from the circulation toward an extravascular inflammatory insult. Following shear-resistant adhesion to the vessel wall, L-selectin has frequently been reported to be rapidly cleaved from the plasma membrane (known as ectodomain shedding), with little knowledge of the timing or functional consequence of this event. Using advanced imaging techniques, we observe L-selectin shedding occurring exclusively as primary human monocytes actively engage in transendothelial migration (TEM). Moreover, the shedding was localized to transmigrating pseudopods within the subendothelial space. By capturing monocytes in midtransmigration, we could monitor the subcellular distribution of L-selectin and better understand how ectodomain shedding might contribute to TEM. Mechanistically, L-selectin loses associationwith calmodulin (CaM; a negative regulator of shedding) specifically within transmigrating pseudopods. In contrast, L-selectin/CaM interaction remained intact in nontransmigrated regions of monocytes. We show phosphorylation of L-selectin at Ser 364 is critical for CaM dissociation, which is also restricted to the transmigrating pseudopod. Pharmacological or genetic inhibition of L-selectin shedding significantly increased pseudopodial extensions in transmigrating monocytes, which potentiated invasive behavior during TEM and prevented the establishment of front/back polarity for directional migration persistence once TEM was complete. We conclude that L-selectin shedding directly regulates polarity in transmigrated monocytes, which affirms an active role for this molecule in driving later stages of the multistep adhesion cascade.

Original language	English
Pages (from-to)	E1461-E1470
Journal	Proceedings of the National Academy of Sciences of the United States of America
Volume	112
Issue number	12
Early online date	9 Mar 2015
DOIs	https://doi.org/10.1073/pnas.1417100112
Publication status	Published - 24 Mar 2015

Keywords

Biglycan
Calmodulin
Invasion
Leukocyte
Migration

UN SDGs

This output contributes to the following UN Sustainable Development Goals (SDGs)

Access to Document

10.1073/pnas.1417100112

L-selectin shedding is activated_IVETIC_Publishedonline9March2015_GOLD VoRFinal published version, 1.65 MB

Cite this

Rzeniewicz, K., Newe, A., Gallardo, A. R., Daviesa, J., Holt, M. R., Patel, A., Charras, G. T., Stramer, B., Molenaar, C., Tedder, T. F., Parsons, M., & Ivetic, A. (2015). L-selectin shedding is activated specifically within transmigrating pseudopods of monocytes to regulate cell polarity in vitro. Proceedings of the National Academy of Sciences of the United States of America, 112(12), E1461-E1470. https://doi.org/10.1073/pnas.1417100112

@article{e2ca978676144fec9f0def539f149e35,

title = "L-selectin shedding is activated specifically within transmigrating pseudopods of monocytes to regulate cell polarity in vitro",

abstract = "L-selectin is a cell adhesion molecule that tethers free-flowing leukocytes from the blood to luminal vessel walls, facilitating the initial stages of their emigration from the circulation toward an extravascular inflammatory insult. Following shear-resistant adhesion to the vessel wall, L-selectin has frequently been reported to be rapidly cleaved from the plasma membrane (known as ectodomain shedding), with little knowledge of the timing or functional consequence of this event. Using advanced imaging techniques, we observe L-selectin shedding occurring exclusively as primary human monocytes actively engage in transendothelial migration (TEM). Moreover, the shedding was localized to transmigrating pseudopods within the subendothelial space. By capturing monocytes in midtransmigration, we could monitor the subcellular distribution of L-selectin and better understand how ectodomain shedding might contribute to TEM. Mechanistically, L-selectin loses associationwith calmodulin (CaM; a negative regulator of shedding) specifically within transmigrating pseudopods. In contrast, L-selectin/CaM interaction remained intact in nontransmigrated regions of monocytes. We show phosphorylation of L-selectin at Ser 364 is critical for CaM dissociation, which is also restricted to the transmigrating pseudopod. Pharmacological or genetic inhibition of L-selectin shedding significantly increased pseudopodial extensions in transmigrating monocytes, which potentiated invasive behavior during TEM and prevented the establishment of front/back polarity for directional migration persistence once TEM was complete. We conclude that L-selectin shedding directly regulates polarity in transmigrated monocytes, which affirms an active role for this molecule in driving later stages of the multistep adhesion cascade.",

keywords = "Biglycan, Calmodulin, Invasion, Leukocyte, Migration",

author = "Karolina Rzeniewicz and Abigail Newe and Gallardo, {Angela Rey} and Jessic Daviesa and Holt, {Mark R.} and Ashish Patel and Charras, {Guillaume T.} and Brian Stramer and Chris Molenaar and Tedder, {Thomas F.} and Maddy Parsons and Aleksandar Ivetic",

year = "2015",

month = mar,

day = "24",

doi = "10.1073/pnas.1417100112",

language = "English",

volume = "112",

pages = "E1461--E1470",

journal = "Proceedings of the National Academy of Sciences of the United States of America",

issn = "0027-8424",

publisher = "National Acad Sciences",

number = "12",

}

Rzeniewicz, K , Newe, A , Gallardo, AR, Daviesa, J, Holt, MR , Patel, A, Charras, GT, Stramer, B, Molenaar, C, Tedder, TF, Parsons, M & Ivetic, A 2015, 'L-selectin shedding is activated specifically within transmigrating pseudopods of monocytes to regulate cell polarity in vitro', Proceedings of the National Academy of Sciences of the United States of America, vol. 112, no. 12, pp. E1461-E1470. https://doi.org/10.1073/pnas.1417100112

L-selectin shedding is activated specifically within transmigrating pseudopods of monocytes to regulate cell polarity in vitro. / Rzeniewicz, Karolina ; Newe, Abigail ; Gallardo, Angela Rey et al.
In: Proceedings of the National Academy of Sciences of the United States of America, Vol. 112, No. 12, 24.03.2015, p. E1461-E1470.

Research output: Contribution to journal › Article › peer-review

TY - JOUR

T1 - L-selectin shedding is activated specifically within transmigrating pseudopods of monocytes to regulate cell polarity in vitro

AU - Rzeniewicz, Karolina

AU - Newe, Abigail

AU - Gallardo, Angela Rey

AU - Daviesa, Jessic

AU - Holt, Mark R.

AU - Patel, Ashish

AU - Charras, Guillaume T.

AU - Stramer, Brian

AU - Molenaar, Chris

AU - Tedder, Thomas F.

AU - Parsons, Maddy

AU - Ivetic, Aleksandar

PY - 2015/3/24

Y1 - 2015/3/24

N2 - L-selectin is a cell adhesion molecule that tethers free-flowing leukocytes from the blood to luminal vessel walls, facilitating the initial stages of their emigration from the circulation toward an extravascular inflammatory insult. Following shear-resistant adhesion to the vessel wall, L-selectin has frequently been reported to be rapidly cleaved from the plasma membrane (known as ectodomain shedding), with little knowledge of the timing or functional consequence of this event. Using advanced imaging techniques, we observe L-selectin shedding occurring exclusively as primary human monocytes actively engage in transendothelial migration (TEM). Moreover, the shedding was localized to transmigrating pseudopods within the subendothelial space. By capturing monocytes in midtransmigration, we could monitor the subcellular distribution of L-selectin and better understand how ectodomain shedding might contribute to TEM. Mechanistically, L-selectin loses associationwith calmodulin (CaM; a negative regulator of shedding) specifically within transmigrating pseudopods. In contrast, L-selectin/CaM interaction remained intact in nontransmigrated regions of monocytes. We show phosphorylation of L-selectin at Ser 364 is critical for CaM dissociation, which is also restricted to the transmigrating pseudopod. Pharmacological or genetic inhibition of L-selectin shedding significantly increased pseudopodial extensions in transmigrating monocytes, which potentiated invasive behavior during TEM and prevented the establishment of front/back polarity for directional migration persistence once TEM was complete. We conclude that L-selectin shedding directly regulates polarity in transmigrated monocytes, which affirms an active role for this molecule in driving later stages of the multistep adhesion cascade.

AB - L-selectin is a cell adhesion molecule that tethers free-flowing leukocytes from the blood to luminal vessel walls, facilitating the initial stages of their emigration from the circulation toward an extravascular inflammatory insult. Following shear-resistant adhesion to the vessel wall, L-selectin has frequently been reported to be rapidly cleaved from the plasma membrane (known as ectodomain shedding), with little knowledge of the timing or functional consequence of this event. Using advanced imaging techniques, we observe L-selectin shedding occurring exclusively as primary human monocytes actively engage in transendothelial migration (TEM). Moreover, the shedding was localized to transmigrating pseudopods within the subendothelial space. By capturing monocytes in midtransmigration, we could monitor the subcellular distribution of L-selectin and better understand how ectodomain shedding might contribute to TEM. Mechanistically, L-selectin loses associationwith calmodulin (CaM; a negative regulator of shedding) specifically within transmigrating pseudopods. In contrast, L-selectin/CaM interaction remained intact in nontransmigrated regions of monocytes. We show phosphorylation of L-selectin at Ser 364 is critical for CaM dissociation, which is also restricted to the transmigrating pseudopod. Pharmacological or genetic inhibition of L-selectin shedding significantly increased pseudopodial extensions in transmigrating monocytes, which potentiated invasive behavior during TEM and prevented the establishment of front/back polarity for directional migration persistence once TEM was complete. We conclude that L-selectin shedding directly regulates polarity in transmigrated monocytes, which affirms an active role for this molecule in driving later stages of the multistep adhesion cascade.

KW - Biglycan

KW - Calmodulin

KW - Invasion

KW - Leukocyte

KW - Migration

UR - http://www.scopus.com/inward/record.url?scp=84925434833&partnerID=8YFLogxK

U2 - 10.1073/pnas.1417100112

DO - 10.1073/pnas.1417100112

M3 - Article

AN - SCOPUS:84925434833

SN - 0027-8424

VL - 112

SP - E1461-E1470

JO - Proceedings of the National Academy of Sciences of the United States of America

JF - Proceedings of the National Academy of Sciences of the United States of America

IS - 12

ER -

L-selectin shedding is activated specifically within transmigrating pseudopods of monocytes to regulate cell polarity in vitro

Abstract

Keywords

UN SDGs

Access to Document

Other files and links

Fingerprint

Cite this