Identification and expression of a new member of the pyrokinin/pban gene family in the sand fly Phlebotomus papatasiby Man-Yeon Choi, Neil D. Sanscrainte, Alden S. Estep, Robert K. Vander Meer, James J. Becnel

Journal of Insect Physiology


Insect Science / Physiology


em pa

Est cal, 2-00 a r t i c l e i n f o

Article history:

Received 27 August 2014

Received in revised form 12 May 2015

Accepted 3 June 2015

Available online 4 June 2015


Phlebotomus papatasi a b s t r a c t functions across orders (reviewed in Rafaeli, 2009). These include (among many others) stimulation of sex pheromone production ed for physiologiy cells of the cenKempe, 19 phageal ga (SEG) and is released into the hemolymph via the corpora ca a neurohemal organ. To date, about 200 PK/PBAN family p have been reported from over 40 species (Choi et al., 2012)

Three genes can potentially produce FXPRL peptides: (1) pk/pban gene usually codes for four or five FXPRL-NH2 (or similar sequence) peptides (diapause hormone (DH), PBAN-like peptides); (2) capa gene usually codes for two periviscerokinin (PVK) peptides (with a C-terminal PRV-NH2) and one DH-like PK (different from the DH found in the PBAN/PK gene); and (3) hugin, found in

Drosophila, is the equivalent to pk/pban which codes for two ⇑ Corresponding authors at: USDA-ARS Horticultural Crops Research Laboratory, 3420 NW Orchard Avenue, Corvallis, OR 97330, USA (M.-Y. Choi). United States

Department of Agriculture, Agriculture Research Service, Center for Medical,

Agricultural and Veterinary Entomology (CMAVE), 1600 SW 23rd Drive, Gainesville,

FL 32608, USA (J.J. Becnel).

E-mail addresses: (M.-Y. Choi), (J.J. Becnel). 1 N.S. and M.C. contributed equally to this work.

Journal of Insect Physiology 79 (2015) 55–62

Contents lists availab

Journal of Insec journal homepage: www.elsioral events during development and reproduction (Gäde and

Goldsworthy, 2003). The pyrokinin (PK)/pheromone biosynthesis activating neuropeptide (PBAN) (=pk/pban) family, a major neuropeptide group found in insects, has been shown to have varying

FXPRL-NH2 (or similar sequence), which is requir cal activity, and are produced in the neurosecretor tral nervous system (CNS) of insects (Raina and

Lepidoptera, PBAN is synthesized in the subeso 0022-1910/Published by Elsevier Ltd.

This is an open access article under the CC BY license ( In nglion rdiaca, eptides .1. Introduction

Insect peptide neurohormones regulate a variety of physiological functions such as fat body homeostasis, feeding, digestion, excretion, circulation, reproduction, metamorphosis, and behavand induction of melanization in lepidopterans (Raina et al., 1989; Matsumoto et al., 1990), hindgut muscle contraction in blattodeans (Holman et al., 1987), and most recently, stimulation of trail pheromone biosynthesis in hymenopterans (Choi and Vander

Meer, 2012). This family of peptides shares a C-terminalPyrokinin



Alternative transcriptionThe major family of neuropeptides (NPs) derived from the pk (pyrokinin)/pban (pheromone biosynthesis activating neuropeptide) gene are defined by a common FXPRL-NH2 or similar sequence at the C-termini.

This family of peptides has been found in all insect groups investigated to date and is implicated in regulating various physiological functions, including pheromone biosynthesis and diapause, but other functions are still largely unknown in specific life stages. Here we identify two isoforms of pk/pban cDNA encoding the PBAN domain from the sand fly Phlebotomus papatasi.

The two pk/pban isoforms have the same sequence except for a 63 nucleotide difference between the long and short forms, and contain no alternative mRNA splicing site. Two NP homologues,

DASGDNGSDSQRTRPPFAPRLamide and SLPFSPRLamide are expected, however, sequence corresponding to the diapause hormone was not found in the P. papatasi pk/pban gene. The PBAN-like amino acid sequence homologue SNKYMTPRL is conserved in the gene, but there is no cleavage site for processing a functional peptide. Characterizing the expression of the isoforms in developmental stages and adults indicates that the short form is differentially transcribed depending on the life stage. The P. papatasi pk/pban gene is the only known pk/pban gene with two transcriptional isoforms and from examination of endoproteolytic cleavage sites is expected to produce fewer peptides than most of the pk/pban genes elucidated to date; only Drosophila melanogaster is simpler with a single NP detected by mass spectroscopy. A phylogenetic analysis showed P. papatasi pk/pban grouped more closely with other nematoceran flies rather than higher flies.

Published by Elsevier Ltd. This is an open access article under the CC BY license ( and expression of a new m gene family in the sand fly Phlebotomus

Man-Yeon Choi a,1,⇑, Neil D. Sanscrainte a,1, Alden S. aUnited States Department of Agriculture, Agriculture Research Service, Center for Medi

FL 32608, USA bNavy Entomology Center of Excellence, Box 43, Naval Air Station, Jacksonville, FL 3221ber of the pyrokinin/pban patasi ep a,b, Robert K. Vander Meer a, James J. Becnel a,⇑

Agricultural and Veterinary Entomology (CMAVE), 1600 SW 23rd Drive, Gainesville, 43, USA le at ScienceDirect t Physiology evier .com/ locate/ j insphys sectPRL-NH2 peptides (Jurenka and Nusawardani, 2011) although subsequent studies have shown only a single functional peptide detected by mass spectroscopy (Baggerman et al., 2002, 2005).

While peptides similar to those derived from capa genes have been found in crustaceans (the whiteleg shrimp Litopenaeus vannamei), chelicerates (the tick Ixodes scapularis, Metaseiulus occidentalis), and Caenorhabditis elegans, to date pk/pban genes and peptides have only been found in insects (Torfs et al., 2001; Lindemans et al., 2009; Neupert et al., 2009).

The first PBAN, a 33 amino acid peptide, was discovered in

Helicoverpa zea (Raina et al., 1989), andwhile the physiological function of most PBANs is largely unknown, the majority of functional work has focused on the regulation of sex pheromone biosynthesis in Lepidoptera. Recently, the fire ant PBAN has been demonstrated to stimulate a trail pheromone production in the Dufour’s gland of ant workers (Choi and Vander Meer, 2012). Explorations of these neuropeptides in Diptera have also revealed much about the genes involved in synthesizing these peptides. Some indications of function have been demonstrated (Zdárek et al., 1997; Verleyen et al., 2004) by showing that the PK peptide accelerates pupation in the flesh fly, Sarcophaga bullata. In mosquitoes, expressions of the pk/pban gene, as well as the PBAN and DH receptor genes, have been determined for all life stages of themosquito Aedes aegypti and both receptors were shown to be functional through expression binding assays (Choi et al., 2013). Hellmich et al. (2014) localized