T lymphocytes export proteasomes by way of microparticles: a possible mechanism for generation of extracellular proteasomes

Abstract

The 20S proteasome is almost exclusively localized within cells. High levels of extracellular proteasomes are also found circulating in the blood plasma of patients suffering from a variety of inflammatory, autoimmune and neoplastic diseases. However, the origin of these proteasomes remained enigmatic. Since the proteome of microparticles, small membrane enclosed vesicles released from cells, was shown to contain proteasomal subunits, we studied whether intact proteasomes are actively released into the extracellular space. Using human primary T lymphocytes stimulated with CaCl2 and the calcium ionophore A23187 to induce membrane blebbing we demonstrate that microparticles contain proteolytically active 20S proteasomes as well as the proteasome activator PA28 and subunits of the 19S proteasome regulator. Furthermore, our experiments reveal that incubation of in vitro generated T lymphocyte-microparticles with sphingomyelinase results in the hydrolysis of the microparticle membranes and subsequent release of proteasomes from the vesicles. Thus, we here show for the first time that functional proteasomes can be exported from activated immune cells by way of microparticles, the dissolution of which may finally lead to the generation of extracellular proteasomes.

Cervicovaginal microbiome dysbiosis is associated with proteome changes related to alterations of the cervicovaginal mucosal barrier

“…For example, inflammation attracts antigen-presenting cells that use (immuno)proteasomes for the breakdown of proteins for presentation in the major histocompatibility complex -I, 35 whereas in vitro studies have shown that T cells excrete proteasomes using exosomes and that this excretion is enhanced when T cells are activated. 36 In contrast, antiproteases were least abundant in the dysbiotic VMB groups. A delicate balance between proteases and antiproteases most likely allows for combating infections, whereas at the same time minimizing tissue damage and inflammation.…”

De Novo Disruption of the Proteasome Regulatory Subunit PSMD12 Causes a Syndromic Neurodevelopmental Disorder

“…(C) Cell extracts from PBMCs derived from control subjects and subject 1 (who harbors the c.367C>T [p.Arg123*] nonsense mutation) were prepared with a detergent-free lysis buffer (TSDG) as previously described. 45,46 20 mg of extract was subsequently exposed to 0.2 mM of the Suc-LLVY-AMC substrate (Bachem) and analyzed on 3%-12% gradient gels (native PAGE, Invitrogen) for chymotrypsin-like activity. Size controls consisted of 1 mg of purified spleen-derived 20S and/or 26S complex, as indicated.…”

“…The increased size of the 20S proteasome complex in these cells could be attributed to its association with additional interacting partners, such as PA28, which is constitutively present in immune cells. 45 Two acquisition times, 0.2 ms (above) and 0.8 ms (below), are shown. (D) 10 mg of whole-cell lysate from PBMCs prepared from control subjects and subject 1 with TSDG buffer were incubated in a final 100 mL volume containing 0.2 mM Suc-LLVY-AMC in quadruplicate and in the presence of 2 mM ATP/DTT for various periods of time, as indicated.…”

Ubiquitin-independent protein degradation in proteasomes

“…Показано, что в ответ на различные воздействия происходит перераспределение протеасом в клетке [159][160][161]. Более того, при различных стрессовых воздействиях, например, при разного рода повреждениях или заболеваниях, включая различные формы рака, существенно увеличивается число внеклеточных 20S протеасом (в сыворотке крови, спинномозговой жидкости, жидкости бронхоальвеолярного лаважа) [162][163][164][165][166][167][168].…”