Novel findings on cellular trafficking and targeting for granule storage of neutrophil elastase, a multifunctional effector molecule of innate immunity

Linda Källquist

Research output: ThesisDoctoral Thesis (compilation)

229 Downloads (Pure)

Abstract

Neutrophil elastase (NE) has important roles in innate immunity, killing pathogens and controlling the immune response; but how NE is targeted to developing granules is not understood. Therefore, the aim of this thesis was to investigate the sorting of NE.

Transfection experiments in a leukemic cell line, which were confirmed in normal hematopoietic cells, showed that a population of proNE was targeted to the plasma membrane and endocytosed. This targeting required an intact carboxy-terminal propeptide. Furthermore, modified proNE was not endocytosed, indicating structural requirements for endocytosis.

An association was demonstrated between the tetraspanin CD63 and proNE upon coexpression in COS cells. Furthermore, depletion of CD63 in a promyelocytic cell line (achieved by RNA interference or a CD63 mutant) caused reduced processing of proNE into mature NE and reduced constitutive secretion of proNE. We therefore propose that CD63 may be a transmembrane linker that facilitates granule targeting of proNE.

Results from a monocytic cell line indicated that the sorting of proNE was a multistage process including trafficking to the cell surface, endocytosis through coated vesicles and possibly lipid rafts, and possible conversion to mature NE in late endosomes. The inhibition of proNE’s activation into mature NE was accompanied by the accumulation of proNE, suggesting a requirement for activation before granule targeting.

This research provided new perspectives on the cellular trafficking of NE. The thesis proposes that granule sorting of proNE is facilitated by a tetraspanin protein serving as a transmembrane linker and transporter. The hypothesis needs further testing in primary cells to acquire additional evidence of the interactions involved.
Original languageEnglish
QualificationDoctor
Awarding Institution
  • Division of Hematology and Transfusion Medicine
Supervisors/Advisors
  • Hansson, Markus, Supervisor
  • Olsson, Inge, Supervisor
  • Tapper, Hans, Supervisor
Award date2009 Mar 27
Publisher
ISBN (Print)978-91-86253-14-1
Publication statusPublished - 2009

Bibliographical note

Defence details

Date: 2009-03-27
Time: 13:00
Place: Segerfalksalen, Wallenberg Neurocentrum, BMC, Sölvegatan 17, Lund

External reviewer(s)

Name: Borregaard, Niels
Title: Professor
Affiliation: Department of Hematology, Rigshospitalet, Copenhagen, Denmark

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Tapper H, Källquist L, Johnsson E, Persson AM, Hansson M, and Olsson I.
Neutrophil elastase sorting involves plasma membrane trafficking requiring the
C-terminal propeptide. Exp Cell Res 312:3471-84, 2006.

*Rosén H, *Källquist L, Hansson M, Olsson I. Neutrophil pro-elastase sorting
involving the plasma membrane is structure dependent. Manuscript.
*equal contribution

Källquist L, Hansson M, Persson AM, Janssen H, Calafat J, Tapper H, Olsson I.
The tetraspanin CD63 is involved in granule targeting of neutrophil elastase.
Blood 112:3444-3454, 2008.

Källquist L, Hansson M, Calafat J, Persson AM, Olsson I. Neutrophil elastase
and proteinase 3 trafficking routes in myelomonocytic cells. Submitted for
publication.

Subject classification (UKÄ)

  • Hematology

Free keywords

  • Hematopoietic cell
  • CD63
  • neutrophil elastase
  • targeting
  • trafficking
  • intracellular sorting
  • neutrophil
  • azurophil granule

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