The mononuclear phagocyte (MP) system plays a fundamental role in both innate and adaptive
immunity. It includes three broad classes of MPs extensively characterized in the mouse: (1) macrophages,
including alveolar macrophages, Langerhans cells, and three distinct subtypes of interstitial macrophages; (2)
tissue-trafficking monocytes; and (3) dendritic cells (DCs), which fall into two main types (DC1 and DC2),
though DC2 can be further subdivided. All these MPs, except AMs and LCs, which are unique to lung and skin,
reside in multiple organs, including the heart, skin, liver, and gut. MP subtypes demonstrate a clear division of
labor during innate and adaptive immunity with little to virtually no functional redundancy, which means that
specific interactions among them are crucial for optimal immune responses against viral, bacterial, and fungal
infections. Currently, however, multiple fundamental gaps for the identification and understanding of how these
MPs function in human organs limit our ability to develop prevention and treatment strategies across diseases.
This project will investigate cross-species and cross-tissue homologies at the cellular, gene expression
and functional levels. We will obtain fresh human and mouse tissue from multiple organs (lung, skin, and their
draining lymph nodes), and employ three broad approaches. First, we will use both bulk RNA sequencing
(RNA-seq) and single-cell RNA sequencing (scRNA-seq) to identify cross-species and cross-tissue homology.
RNA-seq provides sequencing depth (i.e., whole-transcriptome coverage), and scRNA-seq provides the ability
to confirm bulk homologous MP subtypes and examine the heterogeneity within previously defined MP
subtypes. Thus, bioinformatics analyses will identify clusters of homologous MP cell types and align them
across species. Second, for each cluster identified, we will identify genes conserved across species and
tissues, and those that are unique to a given homologous MP subtype, termed marker genes. The results of
these analyses will provide specific genetic markers for human MP subtypes and genetic treatment targets.
Broadly speaking, there are two categories of key marker genes we will functionally investigate: those
conserved in human-mouse MP counterparts that (1) have been well-defined in mice, but not previously
investigated in their human counterparts; and (2) not well-defined or extensively studied in either species.
Third, we will undertake a rigorous functional validation of the key genes identified in human-mouse MP
counterparts. This includes (a) in-vivo murine models with selective depletion of specific genes using
transgenic and conditional knockout (KO) mice; (b) in vitro model systems for human MPs, including assays for
antigen acquisition and processing, cellular interactions, and induction of adaptive immune responses; and (c)
create time-lapse videos with cellular-level microscopy for functional and morphological characterization.