Conidiation in Neurospora crassa: vegetative reproduction by a model fungus

REVIEW Conidia ion in Neu ospo a c assa: ege a i e ep oduc ion by a model ungus Ca men Ruge -He e os 1,2 &Luis M. Co ochano 3 Recei ed: 12 Ma ch 2019 /Re ised: 14 May 2019 /Accep ed: 20 May 2019 #Sp inge Na u e Swi ze land AG 2019 Abs ac Asexual de elopmen , conidia ion, in he ilamen ous ungus Neu ospo a c assa is a simple de elopmen al p ocess ha s a s wi h he g ow h o ae ial hyphae. Then, he o ma ion o cons ic ions and subsequen ma u a ion gi es ise o he ma u e conidia ha a e easily dispe sed by ai cu en s. Conidia ion is egula ed by en i onmen al ac o s such as ligh , ae a ion and nu ien limi a ion, and by he ci cadian clock. Di e en egula o y p o eins ac ing a di e en s ages o conidia ion ha e been desc ibed. The ole o ansc ip ion ac o s such as FL, and componen s o signal ansduc ion pa hways such as he cAMP phosphodies- e ase ACON-2 sugges a complex in e play be ween di e en ial ansc ip ion and signal ansduc ion pa hways. Compa isons be ween he molecula basis o conidia ion in N. c assa and o he ilamen ous ungi will help o iden i y common egula o y elemen s. Keywo ds Asexual de elopmen .Conidia ion .Neu ospo a .Spo ula ion In oduc ion Neu ospo a c assa is an ascomyce e he e o hallic ilamen ous ungus ha g ows as b anched mul inuclea ed hyphae wi h pe o a ed sep a. In na u e, species o Neu ospo a ha e been ound in a wide ange o a eas ha include opical, sub op- ical and empe a e egions, and he ungus is easily de ec ed g owing on he su ace o i e-sco ched ege a ion because o i s abili y o me abolize cellulose and he ac i a ion by hea o he sexual spo es (ascospo es) (Jacobson e al. 2006;Jacobson e al. 2004; Luque e al. 2012; Tu ne e al. 2001). N. c assa has been used as a model o ganism o he esea ch on se e al aspec s o euka yo ic biology, including he mechanism o ecombina ion, genome de ence by RNAi, ci cadian clock egula ion and ligh sensing (Da is and Pe kins 2002; Pe kins and Da is 2000; Roche e al. 2014) The li e cycle o N. c assa includes asexual ep oduc ion and he de elopmen o ege a i e conidia ha a e easy o dispe se. The de elop- men al p ocesses ha lead o conidia ion in N. c assa a e, howe e , e y di e en om hose egula ing conidia ion in Aspe gillus nidulans, despi e bo h being membe s o he Ascomyco a. He e, we e iew ou knowledge o he egula- ion, he gene ics and molecula basis o conidia ion in N. c assa. We p opose ha he simpli ied mode o ege a i e ep oduc ion in N. c assa and ela ed ungi may help o un- de s and asexual de elopmen in o he ascomyce es wi h mo e complex conidia ion pa hways. Mo phological e en s du ing conidia ion in Neu ospo a c assa The e a e wo pa hways o asexual ep oduc ion in N. c assa: mac oconidia ion and mic oconidia ion. Mac oconidia ion (hence o h conidia ion) is induced by he ans e om a liq- uid o an ai in e ace, desicca ion and nu ien deple ion, and is in luenced by se e al en i onmen al ac o s such as ligh and CO 2 le els, and by he ci cadian hy hm (Sp inge 1993). Conidia ion is a e y simple de elopmen al p ocess. The in- duc ion o conidia ion p omo es a change in he di ec ion o g ow h o he ege a i e hyphae ha g ow away om he subs a e, and leads o he o ma ion o a mass o ae ial *Ca men Ruge -He e os c. uge -he e [email protected] g.de 1 Cen e o Molecula Biology o he Uni e si y o Heidelbe g (ZMBH), (DKFZ-ZMBH Alliance), Im Neuenheime Feld 282, 69120 Heidelbe g, Ge many 2 Ge man Cance Resea ch Cen e (DKFZ), Im Neuenheime Feld 280, 69120 Heidelbe g, Ge many 3 Depa amen o de Gené ica, Facul ad de Biología, Uni e sidad de Se illa, Apa ado 1095, 41080 Se illa, Spain In e na ional Mic obiology h ps://doi.o g/10.1007/s10123-019-00085-1 hyphae. Abou 4 h a e conidial induc ion, hyphal g ow h changes om apical elonga ion o apical budding leading o he o ma ion o chains o p oconidia ha a e di ided by mi- no cons ic ions. Budding con inues in p oconidial chains, and majo cons ic ions appea app oxima ely 8 h a e he induc ion o conidia ion. This esul s in he sepa a ion o each p oconidia du ing hei ma u a ion in o ull conidia (Fig. 1A). In e conidial junc ions a e clea ed se e al hou s la e , bu agile connec i e h eads hold conidia oge he un il hey a e dispe sed by wind cu en s (Sp inge 1993; Sp inge and Yano sky 1989). The en i e p ocess o conidia ion akes be- ween 12 and 24 h. Howe e , conidia unde go a biochemical ma u a ion pe iod o se e al days be o e hey a e able o e i- cien ge mina ion (Fig. 1B). The o he asexual ep oduc ion pa hway, mic oconidia ion, is o en obse ed in old cul u es. Mic oconidia a e uninuclea ed spo es mo phologically and de elopmen ally di e en om mac oconidia. I has been shown ha mu an s a ec ed in mac oconidia ion a e no a ec ed in mic oconidia ion, sugges ing ha bo h pa hways a e indepen- den and should ha e ew common egula o y elemen s, i any (Maheshwa i 1999). Mic oconidia eme ge om a p o ube - ance in he ege a i e hyphae which cons ic s un il mic oconidia a e libe a ed. Ex e nally, mic oconidia a e smalle han conidia and do no con ain ca o enoids in hei cell wall. They a e less iable han mac oconidia (Sp inge 1993;Sp inge e al.1992; Sp inge and Yano sky 1989). In addi ion, mic ocycle conidia ion has been obse ed in wild ype isola es o N. c assa, bu he gene ics and mo phological cha ac e iza ion ha e been limi ed (Maheshwa i 1991). The egula ion o conidia ion The p ocess o conidia ion is egula ed by a numbe o en i- onmen al ac o s and endogenous signals ha should be p op- e ly coo dina ed by he ungus in o de o p oduce conidia when en i onmen al condi ions a e app op ia e o ege a i e ep oduc ion. Conidia o ma ion is no mally obse ed when he ungus is exposed o ai , bu ca bon s a a ion can induce conidia ion in subme ged mycelia (Madi e al. 1994; Madi e al. 1997). In N. c assa and o he ungi, de elopmen al cy- cles a e o en ini ia ed by he same en i onmen al s esses ha induce he accumula ion o high le els o ehalose, including ca bon s a a ion, inc eased empe a u e and desicca ion (Li e al. 1997; Sa gen and Kal enbo n 1972; Sp inge and Yano sky 1992; Sun e al. 2011). Mo eo e , he p ocess o de elopmen i sel is conside ed o cause physiological s ess on o ganisms. In ma u e conidia, highe le els o ehalose and s ess esponse p o eins may be equi ed o esis ance and su i al (Hä ke e al. 1998; Rensing and Monne jahn 1998). Thus, ehalose may play a dual ole in he cell, unc- ioning as a ese e ca bohyd a e o u u e hyphal g ow h and Fig. 1 (A) De elopmen al imeline o conidia ion in Neu ospo a c assa. (B) Di e en s ages o conidial de elopmen isualized by scanning elec on mic oscopy (a, b) and luo es- cence mic oscopy (c, d, e, ): his- one H1-GFP o nuclei isuali- za ion and calco luo whi e o cell wall s aining. A ows indica e majo and mino cons ic ions (b), hyphal sep a (a, e) and in e conidial junc ions ( ) In Mic obiol as a s ess p o ec an . The mechanism o induc ion o conidia ion a e he ansi ion o an ai in e phase is no well unde s ood bu i has been p oposed ha he de elopmen o conidia helps he ungus o cope wi h he excess o damaging oxygen adicals (Hansbe g e al. 1993; Pe aza and Hansbe g 2002). Ligh egula es conidia ion (Sp inge 1993), and he egu- la ion by ligh o conidia ion equi es he ac i i y o p o eins WC-1 and WC-2 (Lau e and Russo 1990). WC-1 con ains a zinc inge , a ch omopho e-binding domain (LOV) and PAS domains o p o ein-p o ein in e ac ions (Balla io e al. 1996; C os hwai e e al. 1997). The ch omopho e-binding domain binds he la in FAD, allowing WC-1 o ac as a blue-ligh pho o ecep o (F oehlich e al. 2002; He e al. 2002). The p o ein WC-2 con ains a zinc inge and a PAS domain, and in e ac s wi h WC-1 (Linden and Macino 1997) o o m a Whi e Colla Complex (WCC). This complex, upon ligh ex- posu e, binds ansien ly o he p omo e s o ligh -inducible genes o ac i a e hei ansc ip ion (Belden e al. 2007; F oehlich e al. 2002;HeandLiu2005; Smi h e al. 2010), including he p omo e o l ( lu y), a gene ha encodes a key egula o o conidia ion. Conidia ion inc eases in N. c assa cul u es exposed o ligh sugges ing ha ligh may ac i a e he ansc ip ion o key egula o y genes which indeed accu- mula e a e ligh exposu e as i has been shown o l o csp-1 among o he s (Belden e al. 2007;Chene al.2009; Olmedo e al. 2010a;Olmedoe al.2010b). Conidia ion is egula ed by he endogenous ci cadian clock esul ing in bands o conidia e e y 22.5 h o g ow h in he da k (Dunlap and Lo os 2017). The mechanism by which he clock egula es conidia ion has been cha ac e ized in de ail and is based on a nega i e eedback loop in which he posi i e elemen is he WCC which ac i a es ansc ip ion o q.FRQ is he nega i e elemen o he clock and oge he wi h i s pa ne s he RNA helicase FRH and he casein kinase-1 (CK-1) blocks i s own ansc ip ion as well as he ansc ip ion o he o he WCC- egula ed genes (C os hwai e e al. 1997; Dunlap and Lo os 2004; Hein zen and Liu 2007). The oscil- la o y amoun o FRQ du ing he ci cadian cycle p omo es changes in he abundance, phospho yla ion and ac i i y o he WCC du ing g ow h in he da k (B unne and Scha meie 2006;Dunlap2006; Dunlap and Lo os 2006). Gene ics o conidia ion: egula o y genes and p o eins Gene ics has helped o iden i y some o he key elemen s ha egula e conidia ion in N. c assa. Se e al genes equi ed o conidia ion ha e been desc ibed and hei gene ic in e ac ions ha e been es ablished. S ains wi h mu a ions in aconidia e-2 (acon-2)o lu yoid ( ld) a e blocked in he ansi ion om ilamen ous o budding g ow h. Mu a ions in aconidia e-3 (acon-3)o lu y ( l) allow he p oduc ion o mino cons ic ions, bu de elopmen is blocked and ew majo con- s ic ions be ween p oconidia a e de ec ed. Mu a ions in wo conidial sepa a ion genes (csp-1 and csp-2) p e en he sepa- a ion o he c oss walls in ma u ed conidia o elease ee conidia (Sp inge 1993). O hese genes, a en ion has ocused on he p oduc s o genes l (NCU08726), csp-1 (NCU02713) and csp-2 (NCU06095) ha a e ansc ip ion ac o s ha should ac by egula ing ansc ip ion o genes ha pa ic- ipa e in he de elopmen o conidia. These genes a e all ligh -inducible p o iding a connec ion be ween ligh eg- ula ion and conidia ion (Bailey and Ebbole 1998;Chen e al. 2009; Lamb egh s e al. 2009;Pa ée al.2012; Smi h e al. 2010). The l gene has been cha ac e ized in de ail. The FL p o ein is a 792-amino acid polypep ide con aining a Zn 2 Cys 6 binuclea zinc clus e domain belonging o he Gal4p amily (Bailey and Ebbole 1998). Mu a ions in l block conidia ion a he o ma ion o mino cons ic ions, ew hou s a e he in- duc ion o conidia ion (Sp inge and Yano sky 1989). l mRNA accumula es in ae ial hyphae whe e conidia ion- speci ic genes a e exp essed sugges ing an impo an ole o FL in he exp ession o hose genes (Bailey-Sh ode and Ebbole 2004). Howe e , i has been also ound ha l mRNA accumula es sho ly a e induc ion o conidia ion sug- ges ing an addi ional ole o FL in he o ma ion o ae ial hyphae (Co ea and Bell-Pede sen 2002). The impo ance o FL as one o he majo egula o s o conidia ion in N. c assa elies on he obse a ion o conidial de elopmen when l is o e exp essed in subme ged ege a i e hyphae (Bailey- Sh ode and Ebbole 2004). The o e exp ession o l in ege a- i e hyphae leads o he exp ession o eas (Bailey-Sh ode and Ebbole 2004), he gene o he hyd ophobin odle p o ein loca ed on he su ace o ma u ed conidia (Bell-Pede sen e al. 1992; Lau e e al. 1992). This obse a ion suppo s he desc ibed binding o FL o he eas p omo e (Re ngsam an e al. 2005). O he egula o y genes ha e been ound o be up egula ed when l is o e exp essed in ege a i e mycelia, including he conidia ion-speci ic genes con-6 and con-10 (Re ngsam an e al. 2005) suppo ing he p oposal o FL as a conidia ion-speci ic ansc ip ion ac o . I has been desc ibed ha he aconidial pheno ype o a l mu an can be pa ially supp essed by mu a ion in he gene ib-1, a ansc ip- ion ac o ha is in ol ed in he egula ion o he e oka yon incompa ibili y. This would sugges ha FL may egula e conidia ion h ough he ep ession o VIB-1 (Xiang and Glass 2002). The l gene is di ec ly ac i a ed by ligh h ough he binding o he WCC o a ligh egula o y elemen on i s p omo e loca ed a posi ion −640 om ini ia o ATG (Olmedo e al. 2010a). l mRNA accumula es hy hmically in an ACON-2-dependen manne (Co ea and Bell- Pede sen 2002), suppo ing he p oposal ha he hy hmic p oduc ion o conidial bands equi es he hy hmic accumula- ion o l mRNA. In Mic obiol O he conidia ion mu an s ha e been isola ed and cha ac- e ized. ACON-2 is a cAMP phosphodies e ase, bu i s ole in he egula ion o conidia ion emains o be in es iga ed in de ail (G eenwald e al. 2010), and FLD has been p oposed o be a ansc ip ion ac o (Ca illo e al. 2017;McCluskey e al. 2011). ACON-3 is a p o ein wi hou any iden i ied do- main and is he homologue o MedA, a p o ein ha pa ici- pa es in he egula ion o conidia ion o Aspe gillus nidulans (Chung e al. 2011). I is in e es ing o no e he abundance o ansc ip ion ac o mu an s ha a e blocked in conidia ion. CSP-1 is a ligh -inducible zinc inge ansc ip ion ac o (Lamb egh s e al. 2009; Smi h e al. 2010) and CSP-2 was iden i ied as a ligh - egula ed g ainy head-like ansc ip ion ac o (Colo e al. 2006;Pa ée al.2012). CSP-2 is in ol ed in he de elopmen and emodelling o he cell wall and plays a ole in he ac i a ion o genes ela ed wi h de ence and i ulence (Pa é e al. 2012). A hi d ansc ip ion ac o , FLB-3, has been ecen ly cha ac e ized. I is he N. c assa homologue o FlbC, a ansc ip ion ac o ha pa icipa es in he egula ion o A. nidulans conidia ion. The lb-3 mu an is blocked in conidia ion, shows al e ed sexual de elopmen and is unable o comple e he sexual cycle (Boni e al. 2018). These esul s sugges ha FLB-3 plays a key ole in he an- sc ip ional coo dina ion be ween asexual and sexual ep oduc ion. T ansc ip ional egula ion du ing conidia ion The disco e y ha se e al ansc ip ion ac o s egula e conidia ion sugges ed a key ole in ansc ip ional egula ion du ing conidia ion. In addi ion, se e al genes ha e been ound o be highly exp essed du ing conidia ion (Robe s e al. 1988) al hough hei mu an s do no ha e a clea de elopmen al pheno ype. The con genes o N. c assa a e p e e en ially exp essed du ing conidia ion bu some o hem a e also in- duced by ligh in ege a i e mycelia in a WC-dependen man- ne (Co ochano e al. 1995; Lau e and Russo 1991;Madi e al. 1994). The genes con-10 and con-6 con ain se e al eg- ula o y elemen s in hei p omo e s ha a e esponsible o hei ansc ip ional egula ion by de elopmen , ligh and he ci cadian clock (Co ochano e al. 1995; Lee and Ebbole 1998;Olmedoe al.2010b). Simila complex egula ion has been obse ed o genes con-5 and con-13, since hey a e egula ed by ligh and conidia ion al hough hei p omo e s ha e no been ex ensi ely cha ac e ized. The a ailabili y o he N. c assa genome sequence made possible he c ea ion o a gene knockou mu an collec ion o almos all he genes iden i ied in he genome (Colo e al. 2006; Dunlap e al. 2007). The iable knockou mu an s ha e been analysed and hei de elopmen al pheno ypes ha e been cha ac e ized (Bo ko ich e al. 2004; Ca illo e al. 2017; Colo e al. 2006). Se e al s ains bea ing mu a ions in genes coding o ansc ip ion ac o s ha e shown de ec s in di e en s ages o he N. c assa li e cycle. Mos o hem we e ound o ha e impai ed asexual spo ula ion al hough he co - ela ion be ween he pheno ypes and gene exp ession du ing conidia ion has no been s ablished o mos genes ye (Ca illo e al. 2017). In a ew cases, he ole o ansc ip ional egula o s on conidia ion has been in es iga ed. Mu a ions in he Gal4- like ansc ip ion ac o VAD-5 o he el e p o ein VE- 1 educe he g ow h o ae ial hyphae (Bay am e al. 2008; Sun e al. 2012). The gene chc-1 encodes a p o ein wi h a helix-loop-helix binding domain and he mu an shows enhanced conidia ion, in pa icula unde high CO 2 (Sun e al. 2011). Signal ansduc ion pa hways ha egula e conidia ion The key ole o he cAMP phosphodies e ase ACON-2 in conidia ion indica ed ha o he signal ansduc ion pa hways pa icipa ed in he egula ion o conidia ion in addi ion o he ansc ip ional egula ion o conidia ion genes. Conidia ion in N. c assa is also egula ed by signalling pa hways based on he e o ime ic G p o eins and G p o ein- coupled ecep o s (GPCR). G p o eins play key oles as sig- nalling p o eins in euka yo es whe e hey o m he e o ime s composed o α,βand γsubuni s which a e associa ed wi h he plasma memb ane (Ne es e al. 2002; Won e al. 2012). The Gαsubuni binds GTP and GDP and hyd olyses GTP o GDP, and he Gβand Gγsubuni s o m a dime . In he inac- i e s a e, he Gαsubuni binds GDP and he h ee subuni s a e p esen in a complex in associa ion wi h a GPCR. Ligand binding o he GPCR leads o exchange o GTP o GDP on he Gαp o ein and dissocia ion o he Gαand Gβγ dime . Bo h he Gα-GTP and Gβγ moie ies egula e downs eam e ec o p o eins in a ious sys ems, including ion channels, adenylyl-cyclases, phosphodies e ases and phospholipases. GTP hyd olysis on he Gαsubuni allows he GDP-bound Gα o eassocia e wi h he Gβγ dime and he GPCR a he memb ane, eady o eini ia e he signalling cycle (Li e al. 2007). Neu ospo a has h ee Gαsubuni s (GNA 1–3), one Gβ p o ein (GNB-1), one Gγp o ein (GNG-1) and se e al p e- dic ed GPCRs (Li e al. 2007). GNA-1 was he i s iden i ied G p o ein subuni in ilamen ous ungi, and pa icipa es o- ge he wi h he GPCR GPR-4 and he p o ein kinase A (PKA) in he egula ion o ca bon sou ce-dependen apical g ow h and asexual de elopmen (I ey e al. 1996;I ey e al. 2002;I eye al.1999; Li and Bo ko ich 2006; Tu ne and Bo ko ich 1993). GNA-3 egula es conidia ion ia mod- ula ing a cAMP-dependen pa hway and ac s as a nega i e egula o o conidia ion (Kays and Bo ko ich 2004; Kays e al. 2000). The GNB-1 (Gβ)/GNG-1(Gγ) dime is equi ed o he s abili y o Gαp o eins and ac s as a ep esso o In Mic obiol conidia ion (K ys o o a and Bo ko ich 2005;Yange al. 2002). Recen s udies ha e demons a ed ha RIC-8 ( esis an o inhibi o s o cholines e ase) posi i ely egula es Gαsub- uni s, GNA-1 and GNA-3, and dele ion o ic-8 esul s in de ec s in g ow h and asexual de elopmen . Mu a ions ha ac i a e he GNA-1 and GNA-3 p o eins in he Δ ic-8 back- g ound pa ially supp ess Δ ic-8 pheno ypes. RIC-8 in e ac s wi h GNA-1 and GNA-3 in he yeas wo-hyb id assays and ac s as a GEF (guanine nucleo ide exchange ac o ) o GNA- 1 and GNA-3in i o (W igh e al. 2011). The pheno ypes o mu an s in 36 GPCRs ha e been analysed. Mu a ions in 14 GPCR genes lead o al e a ions in conidia ion, speci ically in he o ma ion o ae ial hyphae. Abou hal o he mu an s showed a educ ion, and hal o he mu an had an inc ease in he amoun o ae ial hyphae. Se e al mu an s had pleio o- pic e ec s on de elopmen sugges ing c oss egula ions be- ween hyphal g ow h, sexual and asexual de elopmen in N. c assa (Cab e a e al. 2015). Mu an s in componen s o o he signal ansduc ion pa h- ways show al e a ions in conidia ion. Mu an s in genes coding o calcium signalling componen s sugges a ole o calcium signalling in conidia ion (Ba man and Tamuli 2017). The GTP-binding cy oskele al p o eins sep ins pa icipa e in cell pola i y and he s ains wi h dele ions in sep in genes show al e a ions in conidia ion, among o he de elopmen al pheno- ypes (Be epiki and Read 2013). In addi ion, kinases pa icipa e in he egula ion o conidia ion. The NDR kinase COT-1 egula es hyphal b anching and in e ac s wi h se e al p o eins ha modula e i s egula o y ac i i ies. The in e ac ing p o eins include MOB2A/B, he a ginine me hyl ans e ase SKB1 and he eg- ula o y subuni s o he p o ein phospha ase 2A. Mu a ions in hese p o eins lead o changes in he egula ion o COT-1 ac i i y and al e a ions in conidia ion, sugges ing ha hey play a egula o y ole in his de elopmen al p ocess (D ash e al. 2010; Feldman e al. 2013; Shomin-Le i and Ya den 2017; Zi e al. 2013). The mu an in he second NDR kinase, DBF-2, shows al e a ions in he g ow h o ae ial hyphae and conidia ion (D ash e al. 2010), and a simila pheno ype is obse ed in he mu an in he pu a i e his idine kinase DCC- 1 (Ba ba-Os ia e al. 2011). Compa ison o conidia ion in N. c assa wi h he asexual p og am o o he ascomyce es The widesp ead occu ence o conidia ion in ascomyce es highligh s he impo ance o his p ocess du ing he e olu ion o his g oup o ungi (Be bee and Taylo 2001). N. c assa and A. nidulans di e ged mo e han 300 million yea s ago (Taylo and Be bee 2006; Taylo and Ellison 2010). Conidia ion in A. nidulans has been in es iga ed in de ail and se e al mu an s a ec ed on conidia ion ha e been cha ac e ized (Pa k and Yu 2012). These mu an s p o ide he ools o add ess ques ions abou he e olu ion o conidia ion. Mo phologically, asexual de elopmen in A. nidulans is signi ican ly di e en om ha in N. c assa.InA. nidulans, conidia ion begins wi h he o - ma ion o a oo cell and is ollowed by successi e eme gence o he s alk, esicle, me ulae, phialides and inally ae ial spo es o conidia (Pa k and Yu 2012). The mas e egula o o conidia ion in A. nidulans is encoded by he gen b lA (b is le). Al hough i has no sequence simila i y wi h l, he ac i a ion o b lA ansc ip ion is su icien o induce conidia ion, and in a simila manne o l,b lA is ac i a ed by ligh in a WC-dependen manne (Mooney and Yage 1990; Ruge -He e os e al. 2011). B lA ac i a es ansc ip ion o ano he egula o y gene, abaA (abacus). Unlike b lA, o e exp ession o abaA does no induce conidia ion bu does cause an a es o ege a i e g ow h and induc ion o genes no mally obse ed du ing conidia ion (Adams e al. 1990;Adamse al.1998). The e a e many o he genes ac - ing ups eam o B lA and whose mu a ion gi es ise o aconidial pheno ypes. Mos o hese genes a e conse ed in he Aspe gilli (Ojeda-Lopez e al. 2018). Some o hese genes a e also ound in he N. c assa genome (Fig. 2)and ha e been ound o complemen he de ec on spo ula ion on he co esponding mu an s in A. nidulans (Table 1) (Chung e al. 2011;Shene al.1998). Despi e he p esence o se e al homologous genes be ween A. nidulans and N. c assa, hei conidia ion pa hways di e in bo h hei egula ion and mo phology, highligh ing he simplici y o he conidia ion p og am in N. c assa (Fig. 2). Conclusion Conidia ion in N. c assa has a complex egula ion bu is a simple de elopmen al p ocess when compa ed wi h ege- a i e ep oduc ion in o he ilamen ous ungi. The isola- ion o aconidial mu an s and he iden i ica ion o he genes in ol ed showed ha di e en ial ansc ip ion and signal ansduc ion pa hways ha e key oles in conidia ion. The ole o en i onmen al egula ion in conidia ion is less un- de s ood. The ole o ligh on he egula ion o conidia ion by con olling ansc ip ion o l ando he key egula o y genes has been p oposed, bu he connec ion be ween nu- ien sensing and conidia ion, and how he ansi ion om a liquid o an ai in e phase ac i a es conidia ion emain o be in es iga ed in de ail. I is possible ha some o hese en i onmen al signals a e sensed by any o he GPCRs ha ha e been desc ibed in N. c assa.TheN. c assa genome sequence and he collec ion o single mu an s wi h dele- ions in mos o he genes in he genome ha e p o ided a la ge collec ion o mu an s wi h al e ed conidia ion. The la ge numbe o conidia ion mu an s sugges s ha al e ing conidia ion is no e y di icul , bu iden i ying key egula- o s, like FL o ACON-2, equi ed gene ic sc eens and In Mic obiol s ong mu an pheno ypes, no a educ ion o inc ease in conidia ion. Fu u e wo k should ocus on iden i ying new key egula o s o conidia ion pe haps by looking o condi- ional aconidial mu an s. The compa ison o he conidia ion pa hways in N. c assa and A. nidulans shows a ew common ea u es like he egu- la ion by ligh o key ansc ip ional egula o s, and he key ole o ansc ip ional egula ion, bu many di e ences like hose in mo phology and he use o speci ic p o eins in each pa hway. I seems ha he e is no a basic conidia ion pa hway sha ed by all ascomyce es bu di e en de elopmen al solu- ions based on di e en ial ansc ip ion and common signal ansduc ion pa hways o he p oblem o how o de elop co- nidia. Howe e , we expec ha u he cha ac e iza ion o conidia ion in N. c assa and compa ison wi h o he ilamen- ous ungi will help o iden i y common elemen s in he eg- ula ion o ungal de elopmen . Acknowledgemen s Resea ch in he labo a o y o LMC is suppo ed by he Spanish Minis y o Science, Inno a ion and Uni e si ies (BIO2015- 67148-R) and Eu opean Funds (Eu opean Regional De elopmen Fund, ERDF). Compliance wi h e hical s anda ds Con lic o in e es The au ho s decla e ha hey ha e no con lic o in e es . Re e ences Adams TH, Deising H, Timbe lake WE (1990) b lA equi es bo h zinc inge s o induce de elopmen . Mol Cell Biol 10:1815– 1817 Adams TH, Wiese JK, Yu JH (1998) Asexual spo ula ion in Aspe gillus nidulans. Mic obiol Mol Biol Re 62:35–54 Bailey LA, Ebbole DJ (1998) The lu y gene o Neu ospo a c assa en- codes a Gal4p- ype C6 zinc clus e p o ein equi ed o conidial de elopmen . Gene ics 148:1813–1820 Table 1 Homologous genes be ween A. nidulans and N. c assa and he co esponding pheno ype o he knockou mu an s Aspe gillus nidulans Neu ospo a c assa Pheno ype o he N.c assa mu an s gA –– luG NCU04264 Wild ype lbA NCU08319 Wild ype lbB NCU07379, c -5 Wild ype lbC NCU03043, lb-3 Aconidial lbD NCU01312, ca-1 Wild ype lbE NCU05255 Unknown adA NCU06493, gna-1 Wild ype pkaA NCU06240, pkac-1 Pleio opic s uA NCU01414, asm-1 Sho ae ial hyphae osA NCU05964, os-1 Reduced conidia ion medA NCU07617, acon-3 Aconidial b lA –– abaA –– we A NCU01033 Wild ype –NCU8726, l Aconidial –NCU9739, ld Aconidial AN1251 NCU2713, csp-1 Conidial sepa a ion de ec i e AN4878 NCU06095, csp-2 Conidial sepa a ion de ec i e pdeB NCU00478, acon-2 Aconidial Fig. 2 Compa ison be ween he egula o y genes con olling conidia ion in A. nidulans (in g ey) s. N. c assa (in blue). Conidia ion in A. nidulans is con olled by he mas e egula o b lA which is necessa y and su icien o induce asexual de elopmen . 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