Science - USA (2020-06-05)

inability of the sperm to transit the UTJ (Fig. 3,
F to K) and bind to the ZP (Fig. 3, L and M). By
contrast,Adam28KO males were found to
be fertile (fig. S9). Consistent with our model
that OVCH2 is downstream of NELL2,Ovch2
was not only epididymis-specific (fig. S7),
OVCH2 protein was exquisitely localized to
the IS of the caput epididymis (Fig. 4A), most
proximal to the testis. These results indicate
that epididymal-specific and secreted pro-
tease OVCH2 is required for sperm ADAM3
processing and consequential sperm fertiliz-
ing ability.
This study indicates the presence of a
lumicrine signal pathway in which a testis-
secreted protein (NELL2) signals through a
ROS1-pathway to regulate the epididymal IS
maturation and subsequent secretion of an
epididymis protease (OVCH2) that processes
ADAM3 for sperm fertilizing ability (Fig. 4, B
to D). NELL2 is produced by developing germ
cells in testis and transits through the luminal
spacetotheISoftheepididymistotriggerits
differentiation. The differentiated IS then
secretes many proteins, including proteases
(such as OVCH2) that act on sperm ADAM3 to
make spermatozoa fully functional. Because
various genes are down-regulated inNell2
KO caput epididymis (fig. S7), it is very likely
that epididymal luminal factors other than
proteases also regulate sperm maturation in a
different way but downstream of the NELL2-
ROS1 lumicrine system. Likewise, because

OVCH2 is required for male fertility, OVCH2

may process other sperm and/or epididymis

proteins required for sperm maturation. The

lumicrine concept that we have demonstrated

here in the male reproductive tract may be

applied more broadly to other organs in which

luminal flow occurs.

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ACKNOWLEDGMENTS

We thank the Biotechnology Research and Development (nonprofit

organization) and Department of Experimental Genome Research

for the generation of the mutant mice. We thank T. Enomoto

for sharing resources and advice.Funding:This work was

supported by KAKENHI grants JP15K06999 (to D.K.); JP18K14612

(to T.N.); JP16KK0180 (to Y.F.); and JP25112007, JP17H01394,

JP19H05750 (to M.I.); AMED JP19 g5010001 and the Takeda

Science Foundation (to M.I.); the Eunice Kennedy Shriver National

Institute of Child Health and Human Development (R01HD088412

and P01HD087157 to M.M.M. and M.I.), and The Bill and

Melinda Gates Foundation (INV-001902 to M.M.M. and M.I.).

Author contributions:D.K., M.Ok., M.M.M., and M.I. designed

experiments. D.K., R.Y., T.T., T.N., T.M., K.S., M.K., T.K., Y.F., M.Oz.,

Z.Y., G.M., and K.M.B. performed experiments. M.H. contributed

resources. D.K., M.Ok., M.M.M., and M.I. wrote the manuscript.

Competing interests:The authors declare no competing interests.

Data and materials availability:RNA-seq data obtained from

WT caput, corpus, and cauda epididymis have been deposited in the

Gene Expression Omnibus (GEO) under accession code GSE138517

(www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE138517).

RNA-seq data obtained from WT andNell2KO caput epididymis

have been deposited in the GEO under accession code GSE133920

(www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE133920). All

other data needed to evaluate the conclusions in the paper

are present in the paper or the supplementary materials.

SUPPLEMENTARY MATERIALS

science.sciencemag.org/content/368/6495/1132/suppl/DC1

Materials and Methods

Figs. S1 and S9

Tables S1 and S2

References ( 21 – 31 )

View/request a protocol for this paper fromBio-protocol.

8 July 2019; accepted 12 April 2020

10.1126/science.aay5134

Kiyozumiet al.,Science 368 , 1132–1135 (2020) 5 June 2020 4of4

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