Nuclear Leukocyte Immunoglobulin-like Receptor A3 Is Monomeric and Is Involved in Multiple Layers of Regulated Gene Expression and Translation

The leukocyte immunoglobulin-like
receptor A3 (LILRA3) is a soluble
protein primarily expressed by peripheral blood monocytes and is abundant
in sera of healthy donors. Extracellular LILRA3 is anti-inflammatory
and displays neuro-regenerative functions in vitro. However, its intracellular
expression, distribution, and function(s) remain unknown. Using a
combination of high-resolution confocal and super-resolution microscopy,
we identified intracellular expression of native LILRA3 in the nucleus
of peripheral blood monocytes and in vitro-derived macrophages. This
unexpected nuclear localization of LILRA3 was confirmed in LILRA3-GFP-transfected
HEK293T cells. Western blot of proteins fractionated from primary
macrophages and the transfected HEK293T cells confirmed nuclear localization
of the native and expressed LILRA3 proteins. Interestingly, most of
the LILRA3 in the nucleus was in a monomeric form like the biologically
active secreted protein, while that in the other cellular compartments
was in mixed monomeric, dimeric, and oligomeric forms. The predominant
presence of monomeric LILRA3 in the nucleus was independently corroborated
in transfected live HEK293T cells using the number and molecular brightness
(N&B) analysis method. Immunoprecipitation and mass spectrometric
peptide sequencing studies revealed that nuclear LILRA3 co-immunoprecipitated
with several nuclear proteins involved in host protein synthesis machinery
via direct interactions to a key multifunctional RNA-binding protein,
the Ewing sarcoma breakpoint region 1 protein (EWS) (data are available
via ProteomeXchange with identifier PXD024602). The biological significance
of the nuclear expression of LILRA3 and its interaction with these
key proteins remain to be elucidated.