Two Receptor Tyrosine Phosphatases of the LAR Family Are Expressed in the Developing Leech by Specific Central Neurons as well as Select Peripheral Neurons, Muscles, and Other Cells

Abstract

Receptor protein tyrosine phosphatases (rPTPs) are thought to play a crucial role in neuronal development, particularly in pathfinding by growing processes. We have cloned and sequenced two Hirudo medicinalis rPTPs that are homologous to the Drosophila and vertebrate rPTPs of the Leukocyte common antigen-related (LAR) subfamily. These Hirudo rPTPs, HmLAR1 and HmLAR2, are products of different, homologous genes, both containing two tandem intracellular phosphatase domains and ectodomains with three tandem Ig domains and different numbers of tandem fibronectin type III (FIII) domains. They are expressed in distinct patterns during embryogenesis. HmLAR1 mRNA is expressed by a subset of central and peripheral neurons and by several peripheral muscular structures, whereas HmLAR2 mRNA is expressed by a different subset of central neurons and by the peripheral, neuron-like Comb cells. HmLAR1 and HmLAR2 proteins are located on the neurites of central neurons. In addition, HmLAR2 is expressed on the cell body, processes, and growth cones of the Comb cells. Because of their CAM-like ectodomains and homology to proteins known to be involved in pathfinding and because they are expressed by different subsets of neurons, we hypothesize that HmLAR1 and HmLAR2 participate in navigational decisions that distinguish the sets of neurons that express them. Furthermore, we hypothesize that HmLAR2 is also involved in setting up the highly regular array of parallel processes established by the Comb cells. Lastly, we propose that the HmLAR1 ectodomain on peripheral muscle cells plays a role in target recognition via interactions with neuronal receptors, which might include HmLAR1 or HmLAR2.

Possible role of the receptor protein tyrosine phosphatase HmLAR2 in interbranch repulsion in a leech embryonic cell

“…For visualization of HmLAR2, a rat antiserum (Gershon et al, 1998b) was diluted 1:10. Following primary antisera incubation overnight at 4°C, preparations were washed 5-8 h in wash buffer and treated overnight with a horseradish peroxidase (HRP)-conjugated affinity-purified goat anti-rat IgG [Jackson Immunochemicals; diluted 1:200 in 0.5% Triton X PBS (phosphate-buffered saline) with 10% NGS].…”

“…Fc-HmLAR1, however, did appear to label specifically many of the peripheral nerves extending throughout the body wall [arrowheads in Fig. 5(D)], an interesting finding given that many central neurons are known to express HmLAR1 (Gershon et al, 1998b).…”

“…We recently cloned and characterized two LARlike rPTPs, HmLAR1 and HmLAR2, which are expressed by several types of embryonic cells in leeches of the species Hirudo medicinalis (Gershon et al, 1998b). Of the two, HmLAR2 has the more restricted range of expression during development.…”

“…Furthermore, HmLAR2 protein is expressed at easily detectable levels on the surface membranes of the CC processes and growth cones, including their filopodial extensions (Gershon et al, 1998b), suggesting that this receptor phosphatase could help mediate the development of this complex cellular array. That this is indeed the case was shown by injecting antibodies raised against the extracellular domain of the receptor into the intact embryo.…”

The role of a LAR-like receptor tyrosine phosphatase in growth cone collapse and mutual-avoidance by sibling processes

“…Repeated injection of the antiserum over several days caused the processes of the CCs to deviate from their highly regular paths and to make many navigational errors, compared with controls, including the crossing-over of sibling processes (Gershon et al, 1998b). These observations provided the first clue that HmLAR2 may be playing a role in orchestrating the growth of the parallel arrays of nonoverlapping processes.…”